Abstract
AbstractRecent clinical data have identified infant patients with lethal ITPA deficiencies. ITPA is known to modulate ITP concentrations in cells and has a critical function in neural development which is not understood. Polymorphism of the ITPA gene affects outcomes for both ribavirin and thiopurine based therapies and nearly one third of the human population is thought to harbor ITPA polymorphism. In a previous site-directed mutagenesis alanine screen of the ITPA substrate selectivity pocket, we identified the ITPA mutant, E22A, as a gain-of function mutant with enhanced ITP hydrolysis activity. Here we report a rational enzyme engineering experiment to investigate the biochemical properties of position 22 ITPA mutants and find that the E22D ITPA has two- and four-fold improved substrate selectivity for ITP over the canonical purine triphosphates ATP and GTP, respectively, while maintaining biological activity. The novel E22D ITPA should be considered as a platform for further development of ITPA therapies.
Funder
National Institute of General Medical Sciences of the National Institutes of Health
Publisher
Springer Science and Business Media LLC
Subject
Organic Chemistry,Biochemistry,Bioengineering,Analytical Chemistry