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Molecular structure of Escherichia coli PurT-encoded glycinamide ribonucleotide transformylase.

Thoden JB, Firestine S, Nixon A, Benkovic SJ, Holden HM
Biochemistry 39 8791-802 (2000)
Cited: 27 times
EuropePMC logo PMID: 10913290

Abstract

In Escherichia coli, the PurT-encoded glycinamide ribonucleotide transformylase, or PurT transformylase, catalyzes an alternative formylation of glycinamide ribonucleotide (GAR) in the de novo pathway for purine biosynthesis. On the basis of amino acid sequence analyses, it is known that the PurT transformylase belongs to the ATP-grasp superfamily of proteins. The common theme among members of this superfamily is a catalytic reaction mechanism that requires ATP and proceeds through an acyl phosphate intermediate. All of the enzymes belonging to the ATP-grasp superfamily are composed of three structural motifs, termed the A-, B-, and C-domains, and in each case, the ATP is wedged between the B- and C-domains. Here we describe two high-resolution X-ray crystallographic structures of PurT transformylase from E. coli: one form complexed with the nonhydrolyzable ATP analogue AMPPNP and the second with bound AMPPNP and GAR. The latter structure is of special significance because it represents the first ternary complex to be determined for a member of the ATP-grasp superfamily involved in purine biosynthesis and as such provides new information about the active site region involved in ribonucleotide binding. Specifically in PurT transformylase, the GAR substrate is anchored to the protein via Glu 82, Asp 286, Lys 355, Arg 362, and Arg 363. Key amino acid side chains involved in binding the AMPPNP to the enzyme include Arg 114, Lys 155, Glu 195, Glu 203, and Glu 267. Strikingly, the amino group of GAR that is formylated during the reaction lies at 2.8 A from one of the gamma-phosphoryl oxygens of the AMPPNP.

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  7. Structural studies of tri-functional human GART. Welin M, Grossmann JG, Flodin S, Nyman T, Stenmark P, Trésaugues L, Kotenyova T, Johansson I, Nordlund P, Lehtiö L. Nucleic Acids Res 38 7308-7319 (2010)
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  10. Crystal structure of a lysine biosynthesis enzyme, LysX, from Thermus thermophilus HB8. Sakai H, Vassylyeva MN, Matsuura T, Sekine Si, Gotoh K, Nishiyama M, Terada T, Shirouzu M, Kuramitsu S, Vassylyev DG, Yokoyama S. J Mol Biol 332 729-740 (2003)
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  20. Structural characterization of glycinamide-RNase-transformylase T from Mycobacterium tuberculosis. Chen C, Liu Z, Liu L, Wang J, Jin Q. Emerg Microbes Infect 9 58-66 (2020)
  21. Structural features of Cryptococcus neoformans bifunctional GAR/AIR synthetase may present novel antifungal drug targets. Chua SMH, Wizrah MSI, Luo Z, Lim BYJ, Kappler U, Kobe B, Fraser JA. J Biol Chem 297 101091 (2021)


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