Thiopurine S-methyltransferase pharmacogenetics: functional characterization of a novel rapidly degraded variant allozyme

Abstract

A novel human thiopurine S-methyltransferase (TPMT) variant allele, (319 T>G, 107Tyr>Asp, *27), was identified in a Thai renal transplantation recipient with reduced erythrocyte TPMT activity. The TPMT*27 variant allozyme showed a striking decrease in both immunoreactive protein level and enzyme activity after transient expression in a mammalian cell line. We set out to explore the mechanism(s) responsible for decreased expression of this novel variant of an important drug-metabolizing enzyme. We observed accelerated degradation of TPMT*27 protein in a rabbit reticulocyte lysate. TPMT*27 degradation was slowed by proteasome inhibition and involved chaperone proteins-similar to observations with regard to the degradation of the common TPMT*3A variant allozyme. TPMT*27 aggresome formation was also observed in transfected mammalian cells after proteasome inhibition. Inhibition of autophagy also decreased TPMT*27 degradation. Finally, structural analysis and molecular dynamics simulation indicated that TPMT*27 was less stable than was the wild type TPMT allozyme. In summary, TPMT*27 serves to illustrate the potential importance of protein degradation - both proteasome and autophagy-mediated degradation - for the pharmacogenetic effects of nonsynonymous SNPs.