STRUCTURES OF FREE AND COMPLEXED FORMS OF ESCHERICHIA-COLI XANTHINE-GUANINE PHOSPHORIBOSYLTRANSFERASE

Structures of free, substrate-bound and product-bound forms of Escheri chia coli xanthine-guanine phosphoribosyltransferase (XGPRT) have been determined by X-ray crystallography. These are compared with the prev iously determined structure of magnesium and sulphate-bound XPRT. The structure of free XGPRT at 2.25 Angstrom resolution confirms the flexi bility of residues in and around a mobile loop identified in other PRT ases and shows that the cis-peptide conformation of Arg37 at the activ e site is maintained in the absence of bound ligands. The structures o f XGPRT complexed with the purine base substrates guanine or xanthine in combination with cPRib-PP, an analog of the second substrate PRib-P P, have been solved to 2.0 Angstrom resolution. In these two structure s the disordered phosphate-binding loop of uncomplexed XGPRT becomes o rdered through interactions with the 5'-phosphate group of cPRib-PP. T he cyclopentane ring of cPRib-PP has the C3 exo pucker conformation, s tabilised by the cPRib-PP-bound Mg2+. The purine base specificity of X GPRT appears to be due to water-mediated interactions between the 2-ex ocyclic groups of guanine or xanthine and side-chains of Glu136 and As p140, as well as the main-chain oxygen atom of Ile135. Asp92, together with Lys115, could help stabilise the N7-protonated tautomer of the i ncoming base and could act as a general base to remove the proton from N7 .when the nucleotide product is formed. The 2.6 Angstrom resolutio n structure of XGPRT complexed with product GMP is similar to the subs trate-bound complexes. However, the ribose ring of GMP is rotated by s imilar to 24 degrees compared with the equivalent ring in cPRib-PP. Th is rotation results in the loss of all interactions between the ribosy l group and the enzyme in the product complex. (C) 1998 Academic Press .