The role for glutamic acid at position 196 in human hypoxanthine phosphoribosyltransferase (HPRT) as investigated using site-directed mutagenesis.
The crystal structure of human HPRT reveals the involvement of E196 side chain at the A-B dimer interface. Interference by valine substitution at this position (E196V), as identified in patients with Lesch-Nyhan disease, nearly abolishes enzymatic activity. Kinetic analysis of the active mutants (E196A, E196D, E196Q, and E196R) suggests that interaction between K68 and E196 side chains contributes to stabilization of cis-configuration during the catalytic cycle. The study also provides further insight into the role of A-B dimer interactions relating to K68 in the regulation of cis-trans isomerization that potentially governs the rate-limiting steps in the HPRT reaction.
Canyuk
B
B
Department of Pharmaceutical Chemistry, Faculty of Pharmaceutical Science, Prince of Songkla University, Songkhla, Thailand. bhutorn.c@psu.ac.th
E-Wan
A
A
Keawwijit
W
W
Nualnoi
T
T
Sirisatean
L
L
Tansakul
P
P
Tanthana
C
C
D002384:Catalysis; D019281:Dimerization; D018698:Glutamic Acid / Q000235:genetics / Q000378:metabolism*; D006157:Guanosine Monophosphate / Q000378:metabolism; D006801:Humans; D007041:Hypoxanthine Phosphoribosyltransferase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020836:Protein Structure, Quaternary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012685:Sepharose / Q000378:metabolism
D050505:Mutant Proteins; D011994:Recombinant Proteins; D018698:Glutamic Acid; D006157:Guanosine Monophosphate; D012685:Sepharose; D007041:Hypoxanthine Phosphoribosyltransferase