Crystal structure analysis of S6 modification protein homologue from Thermus thermophilus HB8


Sakai, H.1, Vassylyeva, N. M.1, Matsuura, T.1, Sekine, S.2, Terada, T.1, Shirouzu, M.1, Kuramitsu, S.2,4, Shibata, T.3, Vassylyev, D. G.2, Yokoyama, S.1,2,5

1Genomic Sciences Center, RIKEN Yokohama Institute, 1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama, Kanagawa 230-0045, Japan
2RIKEN Harima Institute at SPring-8, 1-1-1 Kouto, Mikazuki-cho, Hyogo 679-5148, Japan
3RIKEN, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan
4Osaka Univ., 1-1 Machikaneyama, Toyonaka 560-0043, Osaka, Japan
5Univ. of Tokyo, 7-3-1,Hongo Bunkyo-ku, Tokyo 113-0033, Japan

ISGO International Conference on Structural Genomics 2002 (Berlin, Germany, 2002)

It is known that the ribosome protein S6 from Escherichia coli undergoes an unusual post-translational modification as two to six glutamic acids are added at its C-terminus. It was reported that the rimK gene from E. coli is responsible for the addition of glutamic acid residues. It encodes an S6 modification protein with molecular weight of 31.5kDa.

We have cloned the S6 modification protein homologue (S6M) from Thermus thermophilus HB8 and solved its crystal structure.

The T. thermophilus S6M consists of 280 amino acid residues and has a sequence homology of 34% with that of E. coli. The native crystal belongs to the space group C2 with unit cell parameters a=125, b=51,c=104(Å),β=123° (2mol/asymm.). It diffracts X-ray up to 2Å resolution. Since we could not obtain good heavy atom derivatives, we adopted the MAD method using the SeMet substituted protein. The SeMet derivative crystal belongs to the other space group R3 with unit cell parameters a=b=124, c=94(Å) (2mol/asymm.). Unfortunately, they do not diffract X-ray well, so that we used a technique of cross micro seeding. The crashed micro C2-native crystals were seeded into the crystallization drops containing the SeMet substituted protein. The resultant C2-SeMet crystals give higher X-ray diffraction of 2Å, which is comparable to the original C2-native crystal. By using the C2-SeMet crystal, the crystal structure was successfully solved by the MAD method with the data taken at RIKEN beam line BL44B2 at SPring-8.

The structure of S6M consists of three α+β domains, and has a motif of ATP-grasp fold. In spite of the low sequence homology, the overall topology of S6M is rather similar to that of ATP-dependent enzyme such as glutathione synthetase or D-Ala-D-Ala ligase. The structure and putative function of the T. thermophilus S6M will be discussed.



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MATSUURA Takanori
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