Hypoxanthine guanine phosphoribosyltransferase distorts the purine ring of nucleotide substrates and perturbs the pKa of bound xanthosine monophosphate.

Enzymatic efficiency and structural discrimination of substrates from nonsubstrate analogues are attributed to the precise assembly of binding pockets. Many enzymes have the additional remarkable ability to recognize several substrates. These apparently paradoxical attributes are ascribed to the structural plasticity of proteins. A partially defined active site acquires complementarity upon encountering the substrate and completing the assembly. Human hypoxanthine guanine phosphoribosyltransferase (hHGPRT) catalyzes the phosphoribosylation of guanine and hypoxanthine, while the Plasmodium falciparum HGPRT (PfHGPRT) acts on xanthine as well. Reasons for the observed differences in substrate specificities of the two proteins are not clear. We used ultraviolet resonance Raman spectroscopy to study the complexes of HGPRT with products (IMP, GMP, and XMP), in both organisms, in resonance with the purine nucleobase electronic absorption. This led to selective enhancement of vibrations of the purine ring over those of the sugar-phosphate backbone and protein. Spectra of bound nucleotides show that HGPRT distorts the structure of the nucleotides. The distorted structure resembles that of the deprotonated nucleotide. We find that the two proteins assemble similar active sites for their common substrates. While hHGPRT does not bind XMP, PfHGPRT perturbs the pK(a) of bound XMP. The results were compared with the mutant form of hHGPRT that catalyzed xanthine but failed to perturb the pK(a) of XMP.