Photon Factory Activity Report 2014 #32 (2015) B

BL-10C/2014G162

Structure of βB2-crystallin in solution
Rintaro Inoue* ,Nobuhiro Sato, Yojiro Oba and Masaaki Sugiyama
Research Reactor Institute, Kyoto University, Kumatori, Osaka 590-0494, Japan
crystal structure and that in solution state we calculated
the scattering curve based on 1YTQ. In addition to thechange of Rg high Q region was strongly deviated fromexperimental curve, which reinforces that solution
structure is different from that in crystal. At present wetry to fit the experimentally curve with the aid of MDsimulation, which takes into account the dynamics of twodomains. Referring to the preliminary result Rg isfluctuating with the elapse of time, indicating the domain
motion in solution state. More precise data analysis is
still on the progress.

10I(Q) (a. u.)

1 Introduction
The transparency of eye lens is maintained by theordered arrangement of crystallins, which is a major
component protein in the eye lens proteins. Based onreported reults eye lens crystallin is composed of threetypes of crytsallin: α- (45%), β- (20%) and γ-crystallin(35%). Among the above-described three types ofcrystallins only α-crystallin possesses chaperone
functionality for preventing the formation of abnormal
aggregates in the eye lens. On the other hand β- and γcrystallins are the lens specific attained by exteremly highprotein concentration. With aging these crystallins are
graudually convered from native sate to abnormal
aggregated state, leading to the onset of cataract. In orderto undedatnd the onset of abnormal aggregation we haveto unveil the mechnaim of aggregation process in solution
state, which is relatively similar to in-vivo envrionement.
Among the lens specific it is reported that crystallin βB2crystallin is the dominant one . Hence thorugh theaggregation process of βB2-crystallin we have a chance
to garsp the mechnaim of aggregation process in our lens.
However even for the native state of βB2-crystallin thestructure has not been determinned definitely.
Hence we tired to study the structure of native state ofβB2-crystallin in solutiion state as a first step.

101010-3-4-5-6
20.01

3 Results and Discussion
Fig.1 shows SAXS profile of βB2-crystallin in nativestate. At first the radius of gyration (Rg) was evaluatedform Guinier Analysis as shown by Fig.2. The evaluatedRg value was 22.9Å and compared to the Rg (=24.4Å)
calculated from the crystal structure (1YTQ) solution
structure exhibited compact structure. Referring to thecrystal structure βB2-crystallin is composed of twodomains (refer to the inset of Fig. 1), it is therefore
expected that the two domains are closely packed. Inorder to further gain the structural difference between the4567 8 920.1-1Q (Å )Fig. 1: Scattering profile of βB2-crystallin at room
temperature and solid line corresponds to the simulated
scattering curve based on its crystal structure. Inset
indicates the crystal structure of βB2-crystallin.

I(Q) (a. u.)

2 Experiment
We prepared human recombinant βB2-crystallin
expressed by E. coli. Purified βB2-crystallin wasdialyzed aginst 20 mM Tris/HCl, pH7.8 and 150 mM
NaCl buffer. The concentration of βB2-crystallin was set
to 3.5 mg/ml. The SAXS experiments were perfomedwith a SAXS apparatus (SAXES) instlled at BL-10C of
Photon Factory in Institute of Materials Structure Science,High Energy Accelerator Research Organization (KEK),Tsukuba, Japan at room temperature. The obaserved Xray intensity was corrected for the buffer scattering andtransmission. To avoid the sample damage by X-ray theirradiation time was limited to 10 sec.

30.001

0.5

1.0

1.5
2

2.0-2

2.5

3.0x10-3

Q (Å )

Fig. 2: Gunier plot of I(Q) and solid line correspond tothe result of fit with Guinier function.
* rintaro@rri.kyoto-u.ac.jp