The basic pathway for the biosynthesis of inosine 5'-monophosphate (IMP) in vertebrates requires ten enzymatic conversions. Purine nucleotides are the building blocks of RNA and DNA and enzymes of the purine biosynthetic pathway are targets for anticancer, antiviral and antimicrobial drug discovery. Over the past decade three-dimensional structures for most of the enzymes were worked out, revealing many functional details. However, several unresolved issues remain. There are no structures for FGAM synthetase I (PurL, step 4), a 1400 amino acid protein with multiple domains and for FGAM synthetase II, an 800 amino acid protein requiring two additional gene products, PurS and PurQ. Likewise, there are no structures for FAICAR synthetase II (PurP, step 9) and IMP cyclohydrolase II (PurO, step 10) found in methanogenic archaebacteria. There are several different classes of AIR carboxylase (PurE, step 6) having different mechanisms, but only the structure of the class I enzyme is known. In addition, the proposed acylphosphate intermediates for the ATP-grasp enzymes (PurD, PurT, PurK, PurC and probably PurP) have never been characterized nor have the proposed iminophosphate intermediates for AIR synthetase (PurM, step 5) and PurL. No structures are available for the higher order complexes such as PurK/PurE, PurC/PurE, PurS/PurL/PurQ and PurD/PurM/PurN. The purpose of this proposal is to study the structure and function of purine biosynthetic enzymes and to provide structural information currently missing. The research will focus on enzyme mechanism, protein/protein interactions, substrate channeling and protein evolution.
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