Article info 2017 5 (1)    005  (01)   pp.  12 ~ 23
Title Structure determination of the C-terminal fragment of yeast Ski7 using twinned crystal data
Authors Si Hoon Park 1,+ , Yong-Boo Kuk 1,+ , Ji-Young Lee 1 , Byeong-Cheon Jeong 1 and Hyun Kyu Song 1,2, *
Institutions 1 Department of Life Sciences, Korea University, 145 Anam-ro, Seongbuk-gu, Seoul 02841, Korea, 2 Center for Molecular Dynamics and Spectroscopy, Institute of Basic Science, Seoul 02841, Korea. *Correspondence: hksong@korea.ac.kr + These authors contributed equally to the work.
Abstract The structure determination using twinned crystals is challenging although several algorithms have been developed for detwinning the X-ray data. Our crystal of the C-terminal domain 2 and 3 of Ski7 (Ski7-D2/3), a key part of non-stop mRNA decay has a perfect twin with the twin operator [h, -h-k, -l]. Many different efforts for phasing with multiple anomalous dispersion techniques using selenomethionine substituted wild-type and mutant proteins were not successful and the phases were obtained through the molecular replacement method using recently reported structure of C-terminal GTPase domain of Ski7 from Saccharomyces cerevisiae. The overall structure of Ski7-D2/3 is very similar to that of the corresponding domain of ribosome-associated GTPases including eIF5B, eEF1α, and eRF3. Domains 2 and 3 form a β-barrel structure containing several structurally deviated long connecting loops. Although the linker between domain 2 and 3 is very flexible, the relative orientation between them is virtually the same among all structures, showing that the Ski7-D2/3 does not show major conformational movement upon contacting with G domain.