Crystal Structure of Thiaminase-I from Bacillus thiaminolyticus at 2.0 Å Resolution
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- 作者：Nino Campobasso ; Colleen A. Costello ; Cynthia Kinsland ; Tadhg P. Begley ; and Steven E. Ealick
- 刊名：Biochemistry
- 出版年：1998
- 出版时间：November 10, 1998
- 年：1998
- 卷：37
- 期：45
- 页码：15981 - 15989
- 全文大小：154K
- 年卷期：v.37,no.45(November 10, 1998)
- ISSN：1520-4995
文摘
Thiaminase-I catalyzes the replacement of the thiazole moiety of thiamin with a wide varietyof nucleophiles, such as pyridine, aniline, catechols, quinoline, and cysteine. The crystal structure of theenzyme from Bacillus thiaminolyticus was determined at 2.5 Å resolution by multiple isomorphousreplacement and refined to an R factor of 0.195 (Rfree = 0.272). Two other structures, one native and onecontaining a covalently bound inhibitor, were determined at 2.0 Å resolution by molecular replacementfrom a second crystal form and were refined to R factors of 0.205 and 0.217 (Rfree = 0.255 and 0.263),respectively. The overall structure contains two 
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-type domains separated by a large cleft. At thebase of the cleft lies Cys113, previously identified as a key active site nucleophile. The structure with acovalently bound thiamin analogue, which functions as a mechanism-based inactivating agent, confirmsthe location of the active site. Glu241 appears to function as an active site base to increase thenucleophilicity of Cys113. The mutant Glu241Gln was made and shows no activity. Thiaminase-I showsno sequence identity to other proteins in the sequence databases, but the three-dimensional structure showsvery high structural homology to the periplasmic binding proteins and the transferrins.
/-type domains separated by a large cleft. At thebase of the cleft lies Cys113, previously identified as a key active site nucleophile. The structure with acovalently bound thiamin analogue, which functions as a mechanism-based inactivating agent, confirmsthe location of the active site. Glu241 appears to function as an active site base to increase thenucleophilicity of Cys113. The mutant Glu241Gln was made and shows no activity. Thiaminase-I showsno sequence identity to other proteins in the sequence databases, but the three-dimensional structure showsvery high structural homology to the periplasmic binding proteins and the transferrins.