Structural analyses of a purine biosynthetic enzyme from Mycobacterium tuberculosis reveal a novel bound nucleotide

Jerome Le Nours, Esther Bulloch, Zhening Zhang, David Greenwood, Martin Middleditch, James Dickson, Edward Baker

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Abstract

Enzymes of the de novo purine biosynthetic pathway have been identified as essential for the growth and survival of Mycobacterium tuberculosis (Mtb), and thus have potential for the development of anti-TB drugs. The final two steps of this pathway are carried out by the bifunctional enzyme 5-aminoimidazole-4-carboxamide ribonucleotide transformylase /inosine monophosphate cyclohydrolase (ATIC), also known as PurH. This enzyme has already been the target of anti-cancer drug development. We have determined crystal structures of the Mtb ATIC (Rv0957) both with and without the substrate AICAR, at resolutions of 2.5 A and 2.2 A, respectively. As for other ATIC enzymes, the protein is folded into two domains, the N-terminal domain (residues 1-212) containing the cyclohydrolase active site and the C-terminal domain (residues 222-523) the formyl transferase active site. An adventitiously-bound nucleotide was found in the cyclohydrolase active site in both structures, and was identified by NMR and mass spectral analysis as a novel 5-formyl derivative of an earlier intermediate in the biosynthetic pathway, 4-carboxy-5-aminoimidazole ribonucleotide (CAIR). This result and other studies suggest that this novel nucleotide is a cyclohydrolase inhibitor. The dimer formed by Mtb ATIC is different from those seen for human and avian ATICs, but has a similar 50 A separation of the two active sites of the bifunctional enzyme. Evidence in Mtb ATIC for half-the-sites reactivity in the cyclohydrolase domains can be attributed to ligand-induced movements that propagate across the dimer interface and may be a common feature of ATIC enzymes.
Original languageEnglish
Pages (from-to)40706 - 40716
Number of pages11
JournalThe Journal of Biological Chemistry
Volume286
Issue number47
DOIs
Publication statusPublished - 2011

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