OCARINA Communication 2

OCARINA通信
The OCU Advanced Research Institute for Natural Science and Technology

ject

Pro
l
a
i
c
e
p

S

Establishment of the Artificial Photosynthesis Research Center,a base for collaborative research with industry–The largest mystery solved in photosynthesis to which world attention is drawn–
̶ Introduction to research ̶Daisuke KOSUMI, Specially Appointed Associate Professor“Emission of Excess Energy at the Photosynthetic Antennaeof Purple Bacteria”
Keisuke KAWAKAMI, Specially Appointed Associate Professor“Inhibition Mechanism of the Oxygen-Evolving Reactionin Photosystem II Complex by Iodine Ions”
̶ Reports on activities ̶

OCARINA Seminar
2011 Annual General Meeting
Introduction to Facilities:

ICP/MS (Tomomi SHIMONAKA)
X-ray crystallography system (Ikuko MIYAHARA)̶ Topics ̶Prof. Hideki HASHIMOTO“Finishing the CREST Research”

Prof. Nobuo KAMIYA“Beginning 'Basic S' Research”
Professor Nobuo KAMIYA receives the Asahi Prize!

The OCU Advance Research Institute for Natural Science and Technology

Vol.2

OCARINA Communication 2

Special Project

Establishment of the Artificial Photosynthesis Research Center,a base for collaborative research with industry–The largest mystery solved in photosynthesis to which world attention is drawn–
Beginning in the spring of 2013, the Artificial Photosynthesis Research Center at Osaka City University will
begin in earnest. Building upon the results of photosynthesis research carried out at the OCU Advanced
Research Institute for Natural Science and Technology (OCARINA), we are proceeding forward in an
accelerated fashion with research that is aimed at outlets through collaboration with industry. Regarding that,
here we present a roundtable discussion on the future prospects for photosynthesis and artificial photosynthesis
research between OCU President NISHIZAWA; University Board Member YASUMOTO, who has devoted
ceaseless efforts to establishing the Artificial Photosynthesis Research Center; Professor KINOSHITA, directorof OCARINA; and Professor KAMIYA, who solved the greatest mystery of photosynthesis research.

1

Building on the results of photosynthesis research, towards
research aimed at realizing artificial photosynthesis

research. Beyond that, you will taking up the challenge of artificial
photosynthesis, which has been called the ‘dream technology.’

Professor KINOSHITA: OCU’s Advanced Research Institute for Natural
Science and Technology (OCARINA) is made up of 9 people, 2 regularacademic staff and 7 specially appointed researchers; has obtained
funding from the Japan Science and Technology Agency (JST) as aStrategic Creative Research Promotion Project (CREST), from the Japan
Society for the Promotion of Science (JSPS) as a funded project under
Basic Research S, and research funding from many other sources; andis involved in groundbreaking research. Among our projects, one thathas been cited as a major event is Professor Kamiya’s research resultsin solving the greatest mystery of photosynthesis.

Prof. KAMIYA: Often people tell us that our work is “really filled with

Prof. KAMIYA: Photosynthesis research is a rather well-establishedfield, but there was still one obscured point that was not understood.
Then, in 2011, working together with Professor Shen at Okayama
University, we were able to clear that up. Through our research, thepotential for realizing artificial photosynthesis has been heightened,
and research is now accelerating around the world.
President NISHIZAWA: Professor Kamiya’s research was listed by thejournal Science as one of the top 10 world breakthroughs of the year
2011. It was chosen along with the solar rocket probe Hayabusa, which
is a national project of the Japanese government. So I thought that ouruniversity should put even more effort into the project.
Prof. KINOSHITA: That’s right. As one consequence of the great
ripples in every direction caused by Professor Kamiya’s research
results, it turned into a current that I think led to the establishment
now of this Artificial Photosynthesis Research Center. That is what
has made it possible for the Artificial Photosynthesis Research Center
and OCARINA to collaborate together and in the future develop
photosynthesis and artificial photosynthesis research using the mostadvanced equipment.

dreams,” but I think that already now artificial photosynthesis is nota ‘dream’ but has become a research theme that is something that“somehow we want to make it happen, we must make it happen.”
We will deepen the research through a ‘Joint Research Course’ withenterprises and the university working together closely as a tag team.
Board Member YASUMOTO: In the sense that it’s a utilization of naturalenergy, I want us to start quickly and get at least one step closer
to application. With that in mind, while we are still at the level ofbasic research, I’ve been proposing that we should move forward incollaboration with industry. As a result of that, we’ve been able toget the cooperation of the Osaka City government, and we’ve now
reached the point where we’ve established the Artificial Photosynthesis
Research Center.
Prof. KINOSHITA: We’ve held a lot of roundtable discussions andmeetings with people in the Osaka City government, and yes, they
have great expectations for artificial photosynthesis.
Board Member YASUMOTO: Usually, it’s thought to be alright for basic

research to be conducted at universities, but that’s really not the case.
Artificial photosynthesis is really an enormous technology and is atechnology that in the future has the power to change the world, so
it is important at an early stage to be involved in it in a collaboration
between industry and academia. I myself come from a backgroundin industry, and up until now I’ve had the impression that “there’s noresearch done at the university that is aimed at outlets,” and it was
really not possible to realize an industry-academic collaboration inbasic research. That’s why now, in making it happen, I think there is areally important significance.
Prof. KINOSHITA: What has been called the ‘holy grail’ is something

Boar Member YASUMOTO: I believe this is the first research facility inthat we are now trying to grasp. On that point, I think the people in theworld of industry as well have a lot to look forward to.

Japan to be dedicated to Artificial Photosynthesis. Here at Osaka City
University, we have not only Professor Kamiya, but also Professor
Kinoshita and Professor Hashimoto, all specialists in photosynthesis

Prof. KAMIYA: We are doing basic research, so our job is to elucidate
what it is that happens in photosynthesis that occurs in nature.

The OCU Advance Research Institute for Natural Science and Technology

OCARINA Communication 2

The results of that research, from the standpoint of artificial
photosynthesis, become the ‘blueprints.’ And now, these blueprints
have all been revealed. Therefore, we can work on them all together
from now on, including the world of industry and the world ofgovernment.
Boar Member YASUMOTO: That’s right. From now on, engineering

professors will also have to get involved and work out plans for
how to actually make this pay off. We will put together a joint
research office at the university together with enterprises that haveshown an interest in artificial photosynthesis, and researchers fromthose firms will be based there regularly together with regular staff
researchers form the university. We will act to create such a “Joint
Research Course.” And we can expect that there will be wide requests
from people in private industry to also use the large scale researchequipment and facilities that we will be installing. Looking to the
future, I would like to see us create an open research center that canalso respond to requests from the private sector.

phase. Even within the guidelines of OCARINA, there have beenquestions about “how do we get the word out to society?” but up until
now we haven’t moved away from an academic dissemination of
information. At the Artificial Photosynthesis Research Center, we haveto create an organization with the power to disseminate information,
not only through academic channels, but with an awareness more of
Asia’s society as a whole. Right now, in the field of photosynthesis
research, compared to even South Korea or China, our Japanese
researchers, led by Professor Kamiya, are really out in front.
Prof. KAMIYA: In the basic research, I believe we are now the mostadvanced.
Prof. KINOSHITA: Japan has to have the backbone that comes frombeing the leader of the pack. Not being locally limited, but getting
the word out from Osaka to Asia and further to the world. That
perspective will be absolutely necessary.
President NISHIZAWA: That’s quite true. This kind of research isProf. KINOSHITA: This kind of involvement is something that we

researchers cannot start on our own. I think it’s something where weneed the active assistance of the university president and all of theboard members. It needs to be an organization that can disseminate
our work out the society, to Asia, and to the world as a whole.
President NISHIZAWA: Three years ago, when I became university

president, I set out 3 important items on my agenda. Those were,first, to develop education and research in the field of urban scienceand strengthen our function as a think tank for the city of Osaka;
second, to foster members of society with a high level of professional
specialization; and third, to strengthen our international presence.
Artificial photosynthesis fits in precisely with the first item, thefield of urban science. From my first year serving as president, inconversations with the professors in OCARINA, I thought that
this was probably something the university should promote more.
I believe that strengthening the collaboration with industry, andcarrying basic research disciplines forward into a form that can beutilized by society are also part of the university’s job.
Prof. KINOSHITA: From April onwards, we will be entering a newsomething that can’t be carried out by one lab or by one researcher. In
order to put it into practical application, the power of enterprises will
absolutely be necessary. I think this Artificial Photosynthesis ResearchCenter has come about as the place that will connect them all together.
Looking beyond that, since there are essentially parts of this that canbe applied to the society, there will come a time when this will have tobe dealt with even beyond the framework of this research center. Onceit gets to a certain level, I think this research will spread out and speed
up explosively. Up to that point, we are the ones that have to step up
the process.

Proceeding with the research in cooperation with otheruniversities, aiming towards practical implementation in theyear 2030
Prof. KINOSHITA: Actually, we have proposed a road map for the

Artificial Photosynthesis Research Center to the Osaka Municipal
Assembly that aims at practical implementation by 2030. It’s a road
map in which research begins at maximum speed in 2015 and steps
toward practical implementation begin in 2020. At present we have put

The OCU Advance Research Institute for Natural Science and Technology

2

OCARINA Communication 2

Special Project

profileYoshio YASUMOTO
Board of Directors Member, Osaka City University (in charge of industry collaboration,knowledge resources, and information) / Director, Industry-University Collaboration
Promotion Headquarters / Director, New Industries Creation Research Center
B.S. and M.S. in Electrical Engineering, Kyoto University School of Engineering. Ph.D.
in Engineering. Began working at Matsushita Electric (now Panasonic) Ltd. in 1977.
Served as Director of Matsushita Electric’s Overseas R&D Promotion Center, Board
Member, R&D Area Supervisor, etc. In current position since 2010.

in place a system so that research can proceed quietly step by step inline with the road map, but we will first have to explode in 2015. That’s
coming very soon.
Prof. KAMIYA: Speaking honestly, I thought it was a road mapthat set the hurdles quite high, but research at OCARINA isproceeding substantially and the time is now here when the Artificial
Photosynthesis Research Center can start to move, so I think that “thismight work.”
Board Member YASUMOTO: I too was quite surprised by the road map,3

but I certainly feel that we may be able to make it.
Prof. KINOSHITA: All kinds of economic evaluations have come out

as to why 2030 is such an important year. The thing that will havethe most effect is the fact that the balance between oil production anddemand will collapse. People are putting their hopes in things likeshale gas, but there are still a lot of problems, and so I think that willcertainly be a problem in the year 2030. If we haven’t found a way
forward by that time, there will probably be a great deal of social
unrest at the very least. Also, when spin-off research also appears, thatwas presented in the artificial photosynthesis forum, wasn’t it?
Prof. KAMIYA: The breakdown reaction of water in photosynthesisthat we are researching is a place where weed herbicide is at work. If
the structure of that becomes clear, we will probably be able to makemore effective herbicides. In the breakdown of water in artificial
photosynthesis, if we can come to grips with the place where theoxygen producing reaction is, I think that will certainly give birth toother spin-off technologies and knowledge.

Board Member YASUMOTO: The Artificial Photosynthesis Forumwhich began in September 2012 is the venue for joint activities indisseminating information to the public by Osaka City University andthe Osaka City Urban Industries Promotion Center public foundation.
The other day we had talks by this university’s Professor Umena andProfessor Miyake from Osaka University. After that, time was provided
for them to mix together with people from private industry and havean exchange of views, wasn’t there?
Prof. KINOSHITA: After the research presentations at the venue forexchanges of views, there weren’t just exchanges between us and thefirms, but there was quite an active exchange of views between thevarious firms themselves, weren’t there? Professor Miyake of Osaka
University’s Basic Engineering Department says that he will cooperatein every way with our research. Having opinions from a different
perspective is extremely valuable, so we are quite gratified. I would
also like to see us tie up fully with Osaka Prefectural University who
are strong in general applied research.
President NISHIZAWA: That’s right. I think we should advance joint

research with an eye on the future.
Prof. KINOSHITA: We will proceed with the basic research here
at OCU’s OCARINA, and proceed with the applied research incooperation with Osaka Prefectural University. Additionally,
concerning practical application tied to the world of industry, we
will be proceeding with that at the Artificial Photosynthesis ResearchCenter.

profileIsamu KINOSHITA
Director of Osaka City University’s Advanced Research Institute for Natural Scienceand Technology (OCARINA) / Professor, Graduate School of Natural Sciences
B.S. in Chemistry, Tohoku University School of Natural Sciences. M.S., Graduate Schoolof Natural Sciences, Nagoya University. Ph.D. in Natural Sciences. After serving as
Assistant Professor, Lecturer, and Associate Professor at Osaka City University’s Schoolof natural Sciences, in current position since 2003.

The OCU Advance Research Institute for Natural Science and Technology

OCARINA Communication 2

profileYoshiki NISHIZAWA
President and Board Chairman, Osaka City University
Doctor of Internal Medicine, Osaka City University Medical School. Ph. D. in Medicine.
Served as Assistant Professor, Lecturer, Associate Professor, and Professor at Osaka
City University; Vice Director of OCU Hospital; Dean of Graduate School of Medicine
and Chairman of Department of Medicine; Visiting Professor at Thomas Jefferson
University, USA. In current position since 2010.

Making the Artificial Photosynthesis Research Center
Flourish and Activating Young Researchers
Prof. KINOSHITA: The research that we are involved with, concretely

speaking is hybrid artificial photosynthesis. Building on the fact that
we are at the top level of natural photosynthesis in the world, we are
involved in developing an artificial photosynthesis that combines
both the natural and the artificial together. While there are someadvantages to natural photosynthesis, there are also advantageous
features in the artificial kind, and we are moving along well. Actually,in artificial photosynthesis, there is something called ‘modularization.’
Places where light is gathered, places where water is gathered, andplaces where CO2 is cycled, etc.– developing each of these to a highlevel, and finally combining them all– through doing that, I think thatmany kinds of spin-off technologies will emerge.
Board Member YASUMOTO: Speaking of industrial world, since there

are many different kinds of industrial worlds, I think they should all
be able to participate from their respective positions, whether it be
as plant makers, oil companies, electrical appliance makers, etc. We
still don’t know in which of these practical application will happen
the fastest. The direction that we take will have to be determined by
the progress of the research and based on the views of the various
participants.
President NISHIZAWA: If one thinks about returning the benefits
to society, we have to make joint research with private firms anddevelopment of applied research flourish. Additionally, on the other
hand, in order to cultivate the buds so that the flowers may flourish,
we have to secure human resources in basic research that will become
a firm foundation. At first glance, one might think that these two do

not necessarily go together, but if the flowers can be made to bloom
and flourish in one field, then young people will take an interest andgather, and I think that not only that field but research as a whole canbecome invigorated.
Prof. KINOSHITA: Professor Kamiya, since you were awarded with
the Asahi Prize in 2012 for the work you did with Professor Shen Jianren of Okayama University on “Elucidating the Molecular Structure of
Water Breakdown and Oxygen Production in Photosynthesis,” I guess
you’re in the position of having to disseminate a lot of information,aren’t you?
Prof. KAMIYA: A world in which artificial photosynthesis has actually

been achieved will be a world in which the people living at that time
will take artificial photosynthesis for granted. In order to bring thatabout, I think it is extremely important to keep putting out informationthat will energize young researchers at schools and at private firms.
For that purpose, I think the role that the university plays, or the rolethat OCARINA plays, is extremely large. Already we have received all
kinds of inquiries, and since we know that there are needs, we want to
respond to them as much as we possibly can.
Prof. KINOSHITA: The world that we live in has been made throughphotosynthesis. If energy can no longer be supplied by oil and otherfossil fuels that are the products of ancient photosynthesis, then we
have no choice but to supply it by photosynthesis today. It is not justthe energy problem– photosynthesis is a major key for understanding
the world. Beginning this spring, the Advanced Research Institute for
Natural Sciences and Technology and the Artificial Photosynthesis
Research Center, cooperating together, will move forward developing
research on photosynthesis and artificial photosynthesis.

profileNobuo KAMIYA
Professor, Advanced Research Institute for Natural Science and Technology and OCU
Graduate School of Natural Sciences
B.S., M.S., and Ph.D., in Natural Sciences, Nagoya University. Served as Researcher
and Chief Associate Researcher at the RIKEN Institute, and Director of the Office of
Research Technology at RIKEN Harima Institute’s Large Synchrotron Radiation Facility(SPring-8). In current position since 2005.
2012 Recipient of the Asahi Prize.

The OCU Advance Research Institute for Natural Science and Technology

4

OCARINA Communication 2

Introduction to Research

Emission of Excess Energy at
the Photosynthetic Antennaeof Purple Bacteria

The antennae of photosynthetic purple bacteria were bombarded
with light from a femtosecond pulse laser which can discharge verystrong energy in a very short time, and by recording the electron
excitation state at 10 femtosecond intervals, it was discoveredthat of the total energy that is transmitted from the carotenoids tobacteriochlorophyll, 40% is returned to the carotenoids. The resultsof this research were published in the German academic journal
Angewandte Chemie, International Edition (50, pp. 1097-1100,2011).

5• The Role of Photosynthetic Antennae
The photosynthetic antennae of plants and bacteria are composedof two kinds of pigment molecules, carotenoids and (bacterio-)chlorophyll, and the proteins that surround them. The spatial
arrangement of these pigment molecules and proteins differs
according to the plant or bacteria species, but it is known that theantennae of purple bacteria form a supermolecular aggregate in aring shape. Since the (bacterio-) chlorophyll mainly absorb light at thenear-ultraviolet and near-infrared parts of the spectrum, they cannot
effectively utilize light from the visible spectrum in which the sun’s
rays are strongest. Because of that, the other photosynthetic pigments,
the carotenoids, absorb visible light and transmit that energy to the(bacterio-) chlorophyll at ultra high speed and at high efficiency(light harvesting). This process of energy transmission is carriedout between singlet states of electron excitation of the carotenoids
and the (bacterio-) chlorophyll. The energy that is transmitted to the(bacterio-) chlorophyll moves between the antennae and ultimately
is transported to the reaction centers where it is transformed into
electrochemical energy that can be utilized by organic systems.
• Protective Action from Excess Light
When the photosynthetic body is exposed to excess light, a triplet
state of electron excitation of the (bacterio-) chlorophyll is formed,
and as a result the production of singlet oxygen is promoted whichis harmful to the photosynthetic organism. In order to prevent the
production of singlet oxygen, in the photosynthetic organ, energy
from the electron excitation triplet state of the (bacterio-) chlorophyll
is transmitted to the electron excitation triplet state of the carotenoids,
and by doing this the organism is protected from excess light (photoprotection). However, in antennae of the higher plants which live
close to the earth’s surface where the intensity of the sun’s radiationis comparatively strong, before the singlet electron excitation state of
the chlorophyll that is produced by light that is supplied in excess istransformed to a triplet excitation state, the energy is given off as heatto the outside (non-photochemical quenching, or NPQ). NPQ differsfrom the movement of energy through electron excitation triplet
states as above, and loses the singlet excitation state through a nonradiative process so that energy can be dissipated efficiently from theorganism to the outside. In this way, plants that live in environments
where they are exposed to intense light have developed the functionof protecting the organism from light more than that of collecting
light.
• Reverse Energy Flow between Pigment Molecules in the Antennaeof Purple Photosynthetic Bacteria
Between molecules of carotenoids and (bacterio-) chlorophyll inthe singlet state of electron excitation, when the energy transport
is carried out as below, we have discovered that there is not
only one-way movement of energy from the carotenoids to the(bacterio-) chlorophyll, but as shown in the figure, reverse energy

profileDaisuke KOSUMIOCU Advanced Research Institute for Natural Science and Technology
Specially Appointed Associate Professor
B.S. in Physics, Aoyama Gakuin University Department of Scienceand Engineering, 2002.
M.S in Physics, Tohoku University Graduate School of Sciences, 2007.
Ph. D. in Natural Sciences. Served as Research fellows for the Japan
Society for the Promotion of Sciences (2006–2011). In current position
since April, 2011.
Figure: Outline diagram of energy transfer in the light-harvesting
antennae of the photosynthetic purple bacteria.

movement from the (bacterio-) chlorophyll to the carotenoids is also
possible. For example, in the antennae of purple photosynthetic
bacteria, it is possible for excited energy to move from the singlet
electron excitation state Qx of bacteriochlotophyll to the singlet
state S1 of the carotenoids. Focusing on this point, we attempteda search for the pathway of the reverse energy movement fromthe bacteriochlorophyll to the carotenoids using femtosecond
time analysis measurement. As a result, as shown in the figure, on
the S1 collector antenna of the purple bacterium Rhodospirillum
rubrum, of the energy that the carotenoids absorb and transmit
to the bacteriochlorophyll (43%), we found that 17% of that isreturned back again to the carotenoids. We found that this reverse
energy movement is carried out at the bacteriochlorophyll’s Qx
state and the carotenoid S1 state, and it is both a highly efficient(quantum efficiency of 40%) and extremely rapid (75 femtoseconds)
movement of energy. There has been much research going back anumber of years on energy transfer in the light collector antennae ofthe purple bacterium Rhodospirillum rubrum S1, and it is known
that the energy movement efficiency of 30% from carotenoids tobacteriochlorophyll shows very low resistance. Nonetheless, the factthat 40% of the harvested energy is returned to the carotenoids meansthat the reverse energy movement in the light collector antennae of
Rhodospirillum rubrum S1 is a protection mechanism from light thatis similar in efficiency to the higher plants.
• Towards the Creation of Artificial Photosynthetic Antennae
The factor that determines the efficiency of energy transfer inphotosynthetic antennae is the excited energy movement fromcarotenoids to (bacterio-) chlorophyll. The energy transfer efficiency(30~100%) and absorption wavelength spectrum (450~550 nm)
of the antennae of purple bacteria depends on the combinationof the carotenoids. If the low energy transport efficiency of theantennae which absorb light at long wavelengths (550 nm) as in
the Rhodospirillum rubrum that was used in this research has its
origin in the reverse energy movement from the bacteriochlorophyll
to the carotenoids, by understanding the mechanism in detailand establishing technology to control it, the creation of artificial
photosynthetic antennae that absorb a wider spectrum of light andare highly efficient will become possible.

Figure: Outline diagram of energy transfer in the light collector
antennae of the purple photosynthetic bacterium S1.

The OCU Advance Research Institute for Natural Science and Technology

OCARINA Communication 2

Inhibition Mechanism of the
Oxygen-Evolving Reactionin Photosystem II Complex
by Iodine Ions
Iodine Ions inhibit the Oxygen-Evolving Reaction of Plants andCyanobacteria
The Photosystem II (PSII) complex, along with converting the lightenergy of the sun into chemical energy that can be utilized by plants
and cyanobacteria, is a membrane protein complex that catalyzes
water splitting and produces oxygen molecule. The Mn4CaO5-cluster
catalyzes this reaction, and there are many water molecules and anumber of ions around the Mn4CaO5-cluster. They have the functionof efficiently releasing the protons that are given off by the watersplitting reaction from the Mn4CaO5-cluster to the luminal bulk
phase. The oxygen molecules that are formed by photosynthetic
reaction of PSII are necessary for most of life on earth, however, thefact is that the mechanism of water splitting and oxygen evolution
in PSII is not yet completely understood. Completely understanding
this mechanism and applying those results to the developmentof artificial photosynthesis research is one of our missions as
photosynthesis researchers. Previously studies have shown thatchloride ion (Cl) is an essential cofactor for oxygen evolution of
PSII, and that it is possible to replace Cl- with bromide ion (Br-) andiodine ion (I-) which are homologous elements of Cl-. The oxygen-profileKeisuke KAWAKAMIOCU Advanced Research Institute for Natural Science andTechnology, Specially Appointed Researcher
2005. 3. B.S. in Biology, Okayama University Faculty of Science
2010. 3. Ph.D. in Bioscience, Okayama University Graduate Schoolof Natural Science
2010. 4. Appointed as a Japan Society for the Promotion of Science
Special Researcher in the Bioscience Laboratory of Osaka City
University’s Graduate School of Science
2011. 7. Began serving in current position
Field of Specialization: Plant physiology; X-ray crystallography of
the Photosystem II complex

evolving activity of PSII is not inhibited by substitution of Cl- with
Br-, however, the activity is completely inhibited by substitutionof Cl- with I-. Interestingly, the inhibition mechanism by I - is areversible reaction because the activity inhibited by I- is recovered on
re-substitution of I- by either Cl- or Br- (Figure 1).
Conformational Changes in PSII are caused by Iodine Ions
X-ray crystallography is the most powerful method in order todetermine the 3-dimensional structure of proteins, but its resolution
greatly depends on the qualities of the obtained protein crystals. In
particular, membrane protein complexes like PSII are very unstable,
and it is almost impossible to obtain high quality crystals. Our
research group has been involved with research for many years on
improving the quality of PSII crystals, and we have succeeded indetermining the conditions for obtaining high quality PSII crystals
and elucidating the detailed structure of PSII (Umena et al., Nature,2011). By utilizing these conditions to obtain PSII crystals in which Ihave been replaced and carrying out x-ray crystallography analysis,
it has become possible to understand the fine conformational changes
in PSII caused by I-.
We have confirmed that the conformational change of the aminoacid residue surrounding I- (I-1) and the change of distributions ofwater molecules surrounding I-1 are induced. Moreover, newly I(I-3) is bonded surrounding Mn4CaO5-cluster, and we have actuallyconfirmed the conformational changes in PSII caused by substitution
with I- (Figure 2).
By revealing the inhibition mechanism of the oxygen-evolving
reaction in PSII, this will lead to an understanding of the reaction
mechanism of the Mn4CaO5-cluster, and it can be expected that thisresult will contribute to research on artificial catalysis that mimics the
structure of the Mn4CaO5-cluster.

Figure 1: Effect on the oxygen-evolving activity in PSII at each anion
concentration. Cl- or Br- does not inhibit the activity, however, I- completely
inhibits the activity at high concentration condition.

A

B

Figure 2: (A) Conformational changes near the binding site of Chloride ion (Cl-1) by substitution of Cl- with I-. Change in the arrangement of lysine residue (D2Lys317) is confirmed, and the position of anion (I-1) has changed (red arrow). (B) Conformational changes near the binding site of Chloride ion (Cl-2) by substitutionof Cl- with I-. The position of anion (I-2) has changed (red arrow) and also I- (I-3) has binded. Unit numbers are in angstroms (Å).

The OCU Advance Research Institute for Natural Science and Technology

6

OCARINA Communication 2

Reports on Activities
OCARINA Seminar
At OCU’s Advanced Research Institute for Natural Science and Technology(OCARINA), we invite scientists from Japan and overseas
and put on open lectures on an occasional basis. From this year, a series of ‘major seminar’ of the DISCO Party (see note*) is also held
as the OCARINA seminar.
DISCO Party : *Doctor course students’ incorporated scientific communication party. This is a series of seminar run by the doctoral course students
belongs to OCARINA, which is established this year to promote deep understanding among themselves across the boundaries of research fields. In
addition to the weekly closed seminars for students only, the open seminars will be held with domestic/international invited lecturer(s) as the major
seminer.

1st Meeting [Oct. 27, 2010 Place: Bldg. No. 2, Room 220B]
Guest: Tetsuya OGURA (Visiting Prof., OCARINA)“Tequila, Inulin, and Agave Syrup Made from Agave Plants”

7th Meeting[Oct. 19, 2012 Place: OCU Media Center 1F Cultural Exchange Room]
Guest: Mitsuru FURUSAWA (Founder of Neo-Morgan, Inc. Research Lab)
“Theory of Imbalanced Evolution”

2nd Meeting [Mar. 31, 2011 Place: Bldg. No. 2, Room 220B]
Guest: Hitoshi WATARAI (Invited Prof., Nanoscience Education Research Center,
Osaka Univ.)“Liquid interface nanochemistry for the development of the next generation of green
energy– new analytic principles using the outside and the creation of local analytic
chemistry”

8th Meeting [Oct. 23, 2012 Place: OCU Media Center 1F Cultural Exchange Room]
Guest: Yutaka AMAO (Associate Prof., of Engineering, / JST-PRESTO Researcher)“Creating an artificial photosynthesis system to produce solar fuel and matter”

3rd Meeting [Feb. 17, 2012 Place: 1F Cultural Exchange Room]
Guest: Masahiro YAMASHITA (Prof., of Natural Science,Tohoku Univ.)“Quantum molecular spintronics utilizing monomolecular quantum magnets– which
is better, Nomo or Ichiro?”
4th Meeting [April 28, 2012 Place: Bldg. No. 2, Room 220A]
Guest: Jian-Ping ZHANG (Prof., Dept. of Chemistry, Renmin Univ. China)"Morphological Effect on the Photogeneration of Charge Carriers in P3HT/PCBM
Blend Films""Excitation Dynamics of the Light-Harvesting Complexes of Purple Bacterium
Thermochromatium tepidum"

7

5th Meeting (DISCO party Major Seminar #1)[ July 3, 2012 Place: Bldg. No.2, Room 220B]
Guest: Noritake YASUOKA (Visiting Prof., OCARINA Protein Crystallography)“Making medicines based on the 3-dimensional structure of proteins”
Student: Kiyokazu SANTO (D1, Molecular Design Laboratory)
6th Meeting (DISCO party Major Seminar #2)[Aug. 30, 2012 Place: Bldg. No.2, Room 220B]
Guest: Yoshihito HAYASHI (Associate Prof., Kanazawa Univ. Metallic Oxides)“The chemistry of metal oxide molecules”
Student: Chiasa URAGAMI (D1, Physics of Biological Materials and Crystal GrowthLab)

9th Meeting (DISCO party Major Seminar #3)[Nov. 19, 2012 place:Media Center 1F Cultural Exchange Room]
Guest: John T. KENNIS (Prof.,Vrije Univ. Time-resolved Spectroscopy)“Carotenoids in Photosynthetic Light Harvesting and Photoprotection”
Guest: Anjali PANDIT (Researcher,Vrije Univ. Solid State NMR)“In Shape for Photosynthesis”
Guest: Yoshio TEKI (Prof., OCU, ESR)
“Spin Alignment and Spin Dynamics in Photo-Excited States of � –Radicals”
Student: Satoko HORIBE ( D3, Physics of Biological Materials and Crystal GrowthLab)
Student: Keisuke KAWAMOTO (D1, Molecular Design Lab)

10th Meeting [Dec. 11, 2012 Place: OCU Media Center 1F Cultural Exchange Room]
Guest: Ritsuko FUJII (Specially Appointed Associate Prof., OCARINA/ JST-PRESTO
Researcher)“Photosynthesis of marine seaweeds– revealing the mechanism of their efficient
harvesting of sunlight and its application to artificial photosynthesis”
11th Meeting (DISCO Party Major Seminar #4)[Jan. 24, 2013 Place: Bldg. No.2, Room 220B]
Guest: Masaru TATENO (Prof., Univ. of Hyogo, Theoretical Chemistry)“Theoretical analysis of the biological mechanism that the movement of protons
contributes to”
Guest: Yoshimasa FUKUSHIMA (Specially Appointed Assistant Prof., OCARINA,
Low Temperature Spectroscopy)“Analysis of the photoreaction intermediate by low temperature spectroscopy of theblue light photoreceptor TePixD”
Student: Yoshihiko FURUIKE (D1, Laboratory for Biological Structural Chemistry)The 4th Annual meeting

Fiscal Year 2011 OCARINA Annual General Meeting
Date: March 5-6, 2012
Invited Lecturer• Yasuo INOUE (Prof., Tokyo Metropolitan Univ. /Research Supervisor, JST-PRESTO)• Nobuo KAMIYA (Prof., OCARINA)• Hideki HASHIMOTO (Prof., OCARINA)• Jin-Ren SHEN (Prof., Graduate School of Natural Sciences, Okayama Univ.)• Miwa SUGIURA (Associate Prof., Cell Free Science and Technology ResearchCenter, Ehime Univ.)• Osamu ISHITANI (Prof., Graduate School of Science and Engineering, Tokyo
Institute of Technology)• Hideaki OGATA (Max Planck Institute)• Yoshiki HIGUCHI (Prof., Graduate School of Life Science , Univ. of Hyogo)• Masako KATO (Prof., Graduate School of Sciences, Hokkaido Univ.)• Hiroshi TADAOKA (Institute for Molecular Science)• Mikio HOSHINO (RIKEN)• Shigemasa ARAI (Hill Research, Ltd. CEO)• Yutaka AMAO (Associate Prof., Dept. of Applied Chemistry, Oita Univ.)Symposium Outline
The 4th Annual Symposium was spurred on by the issue that the work of revealing of the oxidation center of PSII was selected by thejournal science as one of the top 10 scientific breakthroughs of 2011. With a focus on the research topic of artificial photosynthesis andits related fields e.g., energy, resources, ecosystem, and all of humanity through the environment, a number of important researchers
from all over Japan were invited to attend this meeting and high-spirited discussions were carried out. Additionally, a debrief meetingfor the final year of the Strategic New Development for the Renewal of the Urban Environment project was also held in tandem, where
the 3 Sub groups’ activities over a four year period was reported.
The OCU Advance Research Institute for Natural Science and Technology

OCARINA Communication 2

Introduction to Facilities: ICP-MS
ICP-MS (Inductively Coupled Plasma Mass Spectrometry) is askillful tool to analyze various isotopes with high sensitivity. ICP(Inductively Coupled Plasma) is the plasma that is produced byionizing argon with induction current in a high frequency induction
coil, and the ICP is used as ion source. Vaporized sample withinargon plasma, is broken down into atoms, which are ionized with>90% efficiency. The ionized atoms are introduced into the mass
spectrometer to determine the concentration of isobars.
ICP-MS has extremely outstanding features as an inorganic mass
spectrometer; 1) it is extremely sensitive, and almost all elements canbe measured in concentrations as low as parts per trillion (ppt) level;2) dynamic range is large, and covering 9 orders of magnitude; 3)
multiple elements can be analyzed at the same time; and 4) not only
quantitative analysis but also qualitative (identification of elements
and isotopes) and simultaneous semi-quantitative analyses can beavailable.
In the system installed in our institute (SPQ9700, Hitachi HighTech Science Cooperation, formarly SII Nano Technology, Inc.),liquid samples are nebulized and introduced into the argon plasma.
Thus, major and trace elements and their isotope of various typesof samples can be analyzed if the sample is appropriately prepared
as water solution; e.g., naturally environmental materials suchas surface and ground waters, soils, rocks and minerals, organic
materials such as foodstuffs, plants and animals, inorganic materials
such as steel, precious metals and semiconductor alloy, highly
purified pharmaceuticals, waste matters, and so on. The ICP-MS is
one of the facilities as regulated methods to analyze environmentallycontrolled materials in waters and soils. ICP-MS is an indispensable

Tomomi SHIMONAKA

facility for the trace element and isotope analyses in the wide rangeof field at present.
Elements in nature exist in numbers of different chemical statesaccording to the environmental conditions. Metal and metalloid
elements change those chemical states in association with redox
condition. Bahaviors and toxicity of the elements also change withthe chemical states. For example, arsenite (AsIII) is more soluble andtoxic than arsenate (AsV), and so does hexavalent chromate (CrVI)
than tetravalent one (CrIV). Thus, the speciation of chemical state ofelements is an important task to evaluate the behaviors and toxicity.
Combined system of HPLC/ICP-MS is skillful for the speciation
analysis. HPLC (High Performance Liquid Chromatography) canseparate the different state of the elements, and ICP-MS is used
as a detector of the separated elements. In our system, HPLC
(SHIMADZU Corporation, LC-6A) is attached to the ICP-MS toanalyze different states of arsenic.
Since installation of
the ICP-MS in 2009, we
have been analyzing
arsenic in natural waters,
soils and rocks, traceand rare earth elements
in groundwaters, rocks
and minerals, and leadand cupper in planttissues.

8

Advanced small molecule CCD crystallography system (VariMax with Saturn)Ikuko MIYAHARASingle crystal structural analysis by X-ray diffraction which
determines directly the 3-D structure of molecules from the
distribution of electron intensity is widely utilized from fields inchemistry that deal with organic compounds and organic metal
complexes to fields in pharmacology that deal with chemicalcompounds that are pharmaceutical candidates, crystals in
different states, and co-crystals. Since there are many cases where
even a very small difference in the structure of molecules can havea determining influence on the molecule’s physical properties
or physiological activity, this method of analysis which makes
determination of structures possible with a high level of reliability
is an extremely powerful means of analysis. The equipment for
single crystal structural analysis that we are now adopting isan X-ray diffraction machine that combines the world’s highest
standard high intensity X-ray beam and high sensitivity CCD
camera, in which additionally the diameter of the beam and thelength of the camera can be changed, making it possible to use itwith very small crystals or crystals that have long lattices.

Microfocus rotating anode X-ray generator(RA-Micro7HFM)

Crystal rotating anode generator type with a maximum effective
power of 1.2kW that uses a molybdenum (Mo) target to measuremost of the diffraction points of small crystals with high precision.

Confocal X-ray optical assembly (Confocal Mirror ‘VariMax Mo’)
This is beam concentrating optical assembly using dual gradedartificial multilayered membrane mirrors that are set at right angles
and can concentrate the x-ray beam three dimensionally. Thesemirrors, the longest in the world, make it possible to obtain a very
high intensity beam that is highly monochromatic.

Advanced 18-bit CCD detector technology (Saturn 724 )The large detection surface area (72 x 72 mm) makes datacollection possible at high resolution even with samples that havelarge lattice constants, the camera length can be adjusted, andit has a high dynamic range (22 bits/pixel), and can be used tomeasure many different kinds of samples. Moreover, by achieving
high sensitivity (180 electron/pixel/photon) and yet with super
low noise (0.2 electron/pixel/sec), it is possible to detect the
weak diffraction points of very small crystals with high precision.
Additionally, by combining the high magnification CCD camera
used for sample observation and absorption correction of thecrystal’s outward shape together with the ¼ χ-shape goniometerand the low temperature device for injection of samples, efficientand highly precise measurement is made possible.
We expect that from now on this equipment will be useful acrossa wide range of research fields in the development superconductors
and molecular magnetism and new functional materials thatrealize organic EL, in the development of inorganic and organic
catalysts, in creating pharmaceuticals based on leading compounds
that include the basic structure of natural materials, and in thedevelopment of artificial photosynthesis that mimics natural
photosynthetic systems.

The OCU Advance Research Institute for Natural Science and Technology

OCARINA Communication 2

Topicsprofile9

Hideki HASHIMOTO

profileOCU Advanced Research Institute for Natural Science andTechnology Professor
M.S. and Ph.D., Kwansei Gakuin University Graduate Schoolof Science. After serving as Assistant Professor in the Osaka City
Univ. Dept. of Engineering and as Associate Professor in the
Shizuoka Univ. Dept. of Engineering, became Full Professor in
the OCU Graduate School of Science. Has served as a project
leader at OCARINA since it was founded in 2008. Since 2010
has been a Specially Appointed Professor at OCARINA whilestill jointly serving in the Graduate School of Science.

Advanced Research Institute for Natural Science andTechnology Professor
Served as Researcher and Chief Associate Researcher at
the RIKEN Institute, and Director of the Office of ResearchTechnology at RIKEN Harima Institute’s Large Synchrotron
Radiation Facility (SPring-8). Has been Professor in the OCU
Graduate School of Science since 2005. Since 2010 has also
been a Specially Appointed Professor at OCARINA.

Nobuo KAMIYA

Finishing the CREST Research

Beginning ‘Basic S’ Research

Our research project entitled “Functional Control of the Primary
Process of Photosynthesis in nano-space” was adopted as a CREST
project by the Japanese Science and Technology Agency for a 5 year
term beginning in October, 2007. The final evaluation of the CREST
research has been completed, and we received a very high evaluation. I would like to take this opportunity to sum up the results of
the CREST research that we have been doing for the last 5 years.
In this research, we artificially fabricated photosynthetic membranes, and using a new spectroscopic method revealed the primary process of photosynthesis that accompanies the transport of
excited energy, with the goal of verifying guidelines for its utilization as an efficient light-harvesting device. As a result, we firmly
established the means for adjusting the artificial light-harvesting
pigment protein complexes; succeeded in obtaining a high resolution atomic force microscopy (AFM) image of the artificial photosynthetic membranes; utilizing femtosecond-coherent spectroscopy,
we clarified the dynamics of vibronic oscillation of photosynthetic
pigments in a unified manner; and by doing this we completely revealed the transportation mechanism of excited energy between the
carotenoids and bacteriochlorophylls that has an important role in
the primary process of photosynthesis. Also, we firmly establisheda unified theory concerning the Stark absorption spectroscopy,
which is extremely effective in revealing the structure and functionof photosynthetic pigments.
In this way, in relation to revealing the operating mechanisms of
the primary reaction in photosynthesis, by accumulating minuteexaminations from both experimental and theoretical aspects, wewere able to produce numerous research results that now lead theworld in this field. The research results have been reported in many
papers published in the world’s top level academic journals (a totalof 124 articles), and we have vigorously presented our findings tothe outside world at both international conferences and conferences
here in Japan. Moreover, since we have received numerous requests
to give invited lectures at international conferences, our group of
research presenters takes pride in the fact that we are leading theworld in this field.
Revealing the function of the artificial pigment protein complexesin photosynthetic antennae and the design synthesis technology
does not stop with understanding the functioning of the natural
world, but also highlights highly valuable spin-off results in scienceand technology, such as the realization of super-photosynthesis thatbreaks through the limits of the natural photosynthesis, solar batteries with increased pigment sensitivity that adapts the technologyof our device, and beyond that direct production of fuels. If thesecan be practically applied, without a doubt we expect that they will
have great social impacts.
Based on this CREST research, the Artificial Photosynthesis Research Center will begin operating in the spring of 2013, and we intend to take up the development of solar fuels, the next-generationenergy, in an industry-university collaboration.

The project funded under Scientific Research Funding Basic
Research (S) for “Revealing the Oxygen Production Mechanism
at Atomic Level Resolution in the Photosystem II Compound in
Photosynthesis” for a 5 year term began in 2012, made up of theseparticipants: Principal researcher: Professor Nobuo KAMIYA(OCARINA), Joint researcher: Professor Tomoyuki YATSUHASHI(OCU Graduate School of Science, Specializes in Molecular Physics,
jointly serves at OCARINA), Collaborating researchers: Specially
Appointed Assoc. Prof. Keisuke KAWAKAMI (OCARINA) andSpecially Appointed Assoc. Prof. Yasushi UMENA (OCARINA).
Most of life on earth is alive due to photosynthesis, andphotosynthesis is one of the most important processes forthe realization of a sustainable society for human beings. The
compound photosystem II (PSII) absorbs sunlight and breaks downwater, and along with molecular oxygen it produces electronsand protons. In April 2011, Kamiya et al. announced that they hadfor the first time in the world succeeded in revealing the detailed
chemical structure of the Mn4CaO5 cluster in PSII that produces
oxygen from water at a resolution of 1.9 Å (Nature [2011] 473,
pp. 55-60). This corresponds to the S1 state of the 5 intermediate
states that explain the oxygen producing reaction (Kok cycle Si
stages, i=0~4), but in this new Basic (S) research our goal is to go
further and conduct X-ray crystal structure analysis of states S0
and S2, and provide the basic information that is necessary fordeveloping catalysts for water reduction and oxygen production.
In this research we will first of all conduct X-ray diffraction
structural analysis of: 1) two kinds of ion substitutions for thechloride ions (Cl-) that contribute to the production of oxygen in
PSII, bromide ions (Br-) and iodide ions (I-); 2) compounds with
5 varieties of phytocides which block movement of electrons thatare connected to oxygen production; and 3) an altered state of PSII
that lacks a subunit of small molecular mass (PsbM) due to genetic
manipulation, for a total of 8 kinds of crystals for X-ray diffraction
structure analysis. In each of these PSII varieties, it is possible

Introducing a New Staff

The OCU Advance Research Institute for Natural Science and Technology

profileYoshimasa FUKUSHIMAOCU Advanced Research Institute for Natural Science andTechnology Specially Appointed Assistant Professor
B.S. in Physics, Nagoya University School of Science. M.S. and Ph.
D., Nagoya Univ. Graduate School of Sciences. After serving as a
researcher at the Nagoya Univ. Graduate School of Sciences andthe same university’s Center for Gene Research, began serving in
his current position in November, 2012.

OCARINA Communication 2that changes will be seen in electron transport activity, and it ispossible the changes will also extend to the Mn4CaO5 clusters.
Particularly in 1) in the iodide substitution, due to the reduction
power of the iodine, there is a high possibility that the S0 state will
be realized. On the other hand, in the Mn4CaO5 cluster, when lightfalls on PSII in state S1, one electron is emitted and it becomes state
S2. The method for creating the S2 state when it comes to liquidsamples of PSII has already been firmly established, but in crystal
samples that are uses for X-ray diffraction structure analysis,
peculiar circumstances arise. That is that because PSII contains a
lot of chlorophyll and carotenoids that absorb visible light, crystals
where PSII is condensed in high concentrations appear to be almost
completely black. In other words, since almost no visible light canpass through the crystal, it is not easy to create an S2 state that ishomogeneously spread throughout the crystal. At that point inthis research, we will hand it over to Professor Yatsuhashi who
has produced many results in research on the ionization of matterby multiple photons, and using infrared light that PSII does notabsorb, and by using those two photons at the same time we will
be able to create the S2 state. Also, by bringing in as a speciallyappointed assistant professor Dr. Yoshimasa FUKUSHIMA, who
is a specialist in spectroscopy (see photo), we hope to succeed in
fixing crystals with a homogeneous S2 state and analyzing theirstructure with X-ray diffraction analysis.

�Professor Nobuo KAMIYA isAwarded the 2012 Asahi Prize!

�

The Advanced Research Institute for Natural Science andTechnology’s Professor Nobuo Kamiya has received the Asahi
Prize for 2012 (joint recipient: Okayama University’s Professor
SHEN Jian-ren). This was due to the high esteem earned by the
results of his research in revealing the molecular structure of the‘manganese cluster’ which is the key to unlocking the mysteryof photosynthesis.
The Asahi Prize was established in 1929 and is awarded toindividuals or groups in a variety of fields in Japan who haveproduced the most outstanding work in the humanities or
natural sciences and have made a considerable contribution to
the development and improvement of culture and society. Manyof the recipients of the Asahi Prize have gone on to win theNobel Prize or other cultural medal awards in later years.
Additionally, Professor Kamiya’s research results were posted
in the online electronic edition of the British academic journal
Nature in April, 2011, and were also chosen as one of the“Top Ten Breakthroughs in Science for 2011” by the American
scientific journal Science.

I feel deeply honored to have been awarded this historicprize. When I look back on the course of my research on
the X-ray crystallography of photosystem II (PSII) andphotosynthesis, the faces of many people come floating up
into my mind such as my mentor during my university
student days, my seniors who started me out in PSII
research, the people who have supported the research over along period of time, the young people who have participated
together in joint research with me, my colleagues and friends
at my current workplace, and my family.
In natural science research, there are parts where it feels
like a small child whimsically playing, and sometimes new
discoveries are born in the midst of that. That needs to becarefully watched over with a long perspective, and in thatsense, I feel once again that I’ve been lucky to have received
many other people’s assistance over such a long time.
From this point on, I would like to move forward doing
research so that we can arrive as quickly as possible at thegoal of completely understanding the water reduction andoxygen producing mechanism of PSII which will lead us to
artificial photosynthesis.
Nobuo KAMIYA, Professor, Advanced Research Institute for
Natural Science and Technology

The OCU Advance Research Institute for Natural Science and Technology

10Osaka City University Advanced Research Institutefor Natural Science and Technology (OCARINA)
OCARINA, the OCU Advanced Research Institute for Natural Science andTechnology, was established as a cross-disciplinary research organization that would
strategically fuse together the knowledge and wisdom of this university to solve the
top priority issues. This institute is meant to be the major player in project research toachieve the goals of “Through a project system, fusing together the most advanced
science and technology across research disciplines, to deal with the foremost complex
issues facing humanity of energy, resources, ecosystems, and the environment on a
global scale, to speak out as scholars and members of society and to nurture humanresources, and to effectively give back the findings that we obtain to the society andthe local community.”
In 2012 we began a new project for “Achieving the production of solar fuels
through artificial photosynthesis.” In the summer of 2012 the construction of a newbuilding for complex of university science has been begun including many floor
spaces for OCARINA. Additionally, the Artificial Photosynthesis Research Center(ReCAP) was in construction as a facility for the industry-university joint researches.
It will open in 2013. In conjunction with that, two new full-time academic staffs will
be joining us from April 2013, and we intend to make our research organization evenmore active.
With the desire to attempt to solve global scale problems as quickly as possible, apart of our facility has already been active moving on ahead, but the upgrading of
all of our facilities is proceeding steadily, and we intend to keep moving forwardmaking this research institute take on the role of base for disseminating world
leading research from Osaka to the world.

Osaka City University Advanced Research Institutefor Natural Science and Technology
3-3-138 Sugimoto, Sumiyoshi-ku
Osaka-shi, 558-8585 JAPAN
Tel: (+81) 06-6605-3634(changed)
Fax: (+81) 06-6605-3636
Midosuji sub.

JR Shinkansen

Shin-Osaka
to Kyoto

to Shin-Kobe

to Kobe

JR Kobe line

Osaka
JR Tozai line

Kyobashi
OSAKA-loop line

Tsuruhasi

Tennoji
Abiko

Sugimoto-cho

Sugimoto
Campus (OCARINA)
Kansai Airport

Hineno
JR Hanwa line

(Original Japanese version published February, 2013)