Steve Ealick's Research Group
Saccharomyces cerevisiae Thiazole Synthase
3FPZ thiazole synthase complexed with adenosine diphospho-5-(β-ethyl)-4-methyl-thiazole-2-carboxylic acid (ADT)
*4Y4L C205S mutant with glycine imine
In contrast to bacteria, thiamin biosynthetic studies in eukaryotes are still at an early stage. Labeling studies in Saccharomyces cerevisiae have demonstrated that the thiamin thiazole is biosynthesized from an unidentified five carbon carbohydrate.The structure of the S. cerevisiae thiazole synthase (ScThi4) was solved by molecular replacement (PDB code1RP0) and reveals the nature of binding to adenosine diphospho-5-(β-ethyl)-4-methyl-thiazole-2-carboxylic acid (ADT) found in the enzyme active site. The identification of this product suggests that NAD is the source of the carbohydrate precursor. Our recent studies* of the structures of the C205S mutant of ScThi4 and the structures of Methanococcus jannaschii thiazole synthase (MjThi4) and Methanococcus igneus thiazole synthase (MiThi4), which lack the active site cysteine, reveal the structural basis for the iron-dependent mechanism of sulfur transfer in archael and yeast thiazole synthases.
The ScThi4 monomer consists of ten β-strands and nine α-helices. Both the N and C terminal helices are amphipathic. ADT is shown bound to the active site.
|Thi4 exists as an octamer with two monomers in the asymmetric unit. The octameric complex may be viewed as a tetramer of dimers with many interdimeric contacts at the interface between two monomers. The outer and inner surfaces are lined with hydrophilic residues.|
Jurgenson CT, Chatterjee A, Begley TP and Ealick SE. Structural insights into the function of the thiamin biosynthetic enzyme Thi4 from Saccharomyces cerevisiae. Biochemistry 45:11061-11070 (2006).
Chatterjee A, Jurgenson CT, Schroeder FC, Ealick SE, and Begley TP. Thiamin Biosynthesis in Eukaryotes: Characterization of the Enzyme-Bound Product of Thiazole Synthase from Saccharomyces cerevisiae and Its Implications in Thiazole Biosynthesis. J. Am. Chem. Soc. 128:7158-7159 (2006).
Chatterjee A, Abeydeera ND, Bale S, Pai PJ, Dorrestein PC, Russell DH, Ealick SE, and Begley TP. Saccharomyces cerevisiae THI4p is a suicide thiamine thiazole synthase. Nature 478:542-6 (2011).
*Zhang X, Eser BE, Chanani PK, Begley TP, and Ealick SE. Structural Basis for Iron-mediated Sulfur Transfer in Archael and Yeast Thiazole Synthases. Biochemistry [Epub ahead of print]:(2016). PubMed
*Eser BE, Zhang X, Chanani PK, Begley TP, and Ealick SE. From suicide enzyme to catalyst: the iron-dependent sulfide transfer in Methanococcus jannaschii thiamin thiazole biosynthesis. J. Am. Chem. Soc. 138:3639-3642 (2016). PubMed