3NL5: The Crystal Structure Of Candida Glabrata Thi6, A Bifunctional Enzyme Involved In Thiamin Biosyhthesis Of Eukaryotes

Citation:
Abstract
THI6 is a bifunctional enzyme found in the thiamin biosynthetic pathway in eukaryotes. The N-terminal domain of THI6 catalyzes the ligation of the thiamin thiazole and pyrimidine moieties to form thiamin phosphate, and the C-terminal domain catalyzes the phosphorylation of 4-methyl-5-hydroxyethylthiazole in a salvage pathway. In prokaryotes, thiamin phosphate synthase and 4-methyl-5-hydroxyethylthiazole kinase are separate gene products. Here we report the first crystal structure of a eukaryotic THI6 along with several complexes that characterize the active sites responsible for the two chemical reactions. THI6 from Candida glabrata is a homohexamer in which the six protomers form a cage-like structure. Each protomer is composed of two domains, which are structurally homologous to their monofunctional bacterial counterparts. Two loop regions not found in the bacterial enzymes provide interactions between the two domains. The structures of different protein-ligand complexes define the thiazole and ATP binding sites of the 4-methyl-5-hydroxyethylthiazole kinase domain and the thiazole phosphate and 4-amino-5-hydroxymethyl-2-methylpyrimidine pyrophosphate binding sites of the thiamin phosphate synthase domain. Our structural studies reveal that the active sites of the two domains are 40 A apart and are not connected by an obvious channel. Biochemical studies show 4-methyl-5-hydroxyethylthiazole phosphate is a substrate for THI6; however, adenosine diphospho-5beta-ethyl-4-methylthiazole-2-carboxylic acid, the product of THI4, is not a substrate for THI6. This suggests that an unidentified enzyme is necessary to produce the substrate for THI6 from the THI4 product.
PDB ID: 3NL5Download
MMDB ID: 86117
PDB Deposition Date: 2010/6/21
Updated in MMDB: 2010/11
Experimental Method:
x-ray diffraction
Resolution: 3.3  Å
Source Organism:
Similar Structures:
Biological Unit for 3NL5: hexameric; determined by author and by software (PISA)
Molecular Components in 3NL5
Label Count Molecule
Proteins (6 molecules)
6
Thiamine Biosynthetic Bifunctional Enzyme
Molecule annotation
Chemicals (18 molecules)
1
6
2
6
3
6
* Click molecule labels to explore molecular sequence information.

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