Steve Ealick's Research Group


Aminoimidazole Ribonucelotide Synthetase (PurM)


PDB file: 1CLI

Description:

The purine biosynthetic pathway in procaryotes enlists eleven enzymes, six of which use ATP. Enzymes 5 and 6 of this pathway, formylglycinamide ribonucleotide (FGAR) amidotransferase (PurL) and aminoimidazole ribonucleotide (AIR) synthetase (PurM) utilize ATP to activate the oxygen of an amide within their substrate toward nucleophilic attack by a nitrogen. AIR synthetase uses the product of PurL, formylglycinamidine ribonucleotide (FGAM) and ATP to make AIR, ADP, and Pi.

The PurM monomer consists of 345 amino acids in a chain that can be divided into two structural domains, which are connected by a well-defined loop. Domain A contains a long, poorly ordered glycine-rich loop, and has four unusually long β-strands. Its first 16 residues, which contain the polyhistidine tag used for purfication, are disordered. The α- helices and β-strands in domain B are shorter than those in domain A.

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The active form of PurM consists of two crystallographically independent dimers, 538 water molecules, and four sulfate ions. The dimer is formed by joining two four-stranded β-sheets (one from each subunit) face to face to form an eight-stranded β-barrel-like structure flanked by eight a helices.

 

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The active site is proposed to be a long groove generated by the interaction of the two monomers. This information is derived from inspecting conserved residues in 23 PurM sequences, from the fact that sulfate co-crystallizes at one end of the cleft, and from affinity-labeling studies.

 

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References:

Li C, Kappock TJ, Stubbe J, Weaver TM and Ealick SE. X-ray crystal structure of aminoimidazole ribonucelotide synthetase (PurM), from the Escherichia coli purine biosynthetic pathway at 2.5 Å resolution. Structure 7:1155-1166 (1999).

Additional work that consists of exploring the ATP binding site of this enzyme is discussed in:

Mueller EJ, Oh S, Kavalerchik E, Kappock TJ, Meyer E, Li C, Ealick SE and Stubbe J. Investigation of the ATP binding site of Escherichia coli aminoimidazole ribonucleotide synthetase using affinity labeling and site-directed mutagenesis. Biochemistry 38:9831-9839 (1999).



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