GTR Interdisciplinary Research Report
Spanish National Cancer Research Centre (CNIO)
Daffa Sean Adinegoro
Affiliation: Laboratory of Molecular Biotechnology, Graduate School of BioagriculturalSciences

Year: D1

Destination: Spanish National Cancer Research Centre (CNIO)
Period: December 2022 – May 2023
Purpose: Expression and purification of anti-GPR54 antibody and protein crystallography
training
Summary:
Using the Ecobody technology that our laboratory invented(Ojima-Kato et al., Scientific Reports, 2017), I successfully
screened some clones of monoclonal antibody against GPR54
receptor protein. For protein characterization, a large amountof protein of both antigen and antibody is required. Besides,for further maximizing the use of anti-GPR54 antibody, I
planned to use it for crystallizing the GPR54 protein which
until now has not been resolved. Since protein crystallization
is not my background, I decided to conduct interdisciplinary
research with Prof. Inès Muñoz from Spanish National Cancer
Research Centre (CNIO). Thus, from December 2022 to May
2023, I went to Spanish National Cancer Research Centre(CNIO) to learn and conduct protein expression and crystallography.
Impression:
Research:Here, I learned to do crystallization using a crystallization robot called Mosquito. For the primary
screening for crystallization condition, there are more than 100 buffer conditions that need to betested. Since dispensing the buffer and protein with more than 100 conditions takes time, theMosquito robot can handle all protein and buffer dispensing in 96 well plate specialized forcrystallization screening. After getting the best buffer condition from primary screening,optimization is done to get more crystal by growing in larger volume of protein. The grown crystal
is then sent to synchrotron to be diffracted by X-ray. Another techniques that I learn is Small
Angle X-ray Scattering (SAXS) where protein does not need to be crystallized and protein
solution is beamed using X-ray. It is much easier compared to standard X-ray crystallography;
however, it has size limitation. Until this report is written, I have learnt the crystallization
technique and helped to crystallize some proteins from other groups in CNIO, but the anti-GPR54
antibody, my project protein, cannot be crystallized due to difficulty of protein purification.
Nevertheless, some conditions to do protein purification has been known, thus improvements ofprotein purification will be done in Japan.
Environment:
Spain in general has lower workload compared to Japan. Here, many people do the jobs (includingexperiment) slowly, not in a rush, since they usually plan the experiment step by step (sometimes
only one step of experiment for one day) until reached their target. Thus, there are some days
when they come to work, but do not do anything since they don’t have things to do on that day.
Meanwhile, during lunch, it is common to have a long lunch time (up to two hours) from 2:00 to4:00 p.m., and then come back to work until 7:00 or 8:00 p.m. Additionally, dinner in Spain
usually starts around 9:00 or 10:00 p.m due to geographical reason where sun sets after 10:00 p.m.
Spain has 40 days paid holiday a year, thus it is common to see many people will take paid holiday
each month. This kind of workstyle makes Spanish people have high life expectancy and do nothave stress in most of their life.
There are many goods Spanish cuisine that I tried when I was in Spain. For example, Paella,Bocadillo, Pollo Asado, etc. It is well known that Spain has the best cuisine in Europe due to theirgeographical location which has more sun and warmer temperature.
Due to Schengen Agreement, it is possible to go to some of European countries without border
restrictions. Flights between Schengen countries is treated as domestic flight and there is noborder check in the land border. During my stay in Spain, I went to Portugal and Andorra by Car
as well as visited my friend who is now studying in Netherlands.
Acknowledgement:
I would like to acknowledge GTR Program for providing me opportunities to do collaborationand interdisciplinary research in Spain. I also would like to acknowledge Prof. Ines G. Munoz,Prof. Hideo Nakano who has allowed me to do this collaboration.