Contrasting behaviors of mutant cystathionine beta-synthase enzymes associated with pyridoxine response

• Published in: Human mutation 2006-05, Vol.27 (5), p.474-482
• Main Authors: Chen, X., Wang, L., Fazlieva, R., Kruger, W.D.
• Format: Article
• Language: English
• Subjects: Alleles; Animals; CBS; cystathionine beta-synthase; Cystathionine beta-Synthase - genetics; Cystathionine beta-Synthase - metabolism; Homocysteine - blood; homocystinuria; Homocystinuria - drug therapy; Homocystinuria - genetics; Humans; Kinetics; Mice; Mice, Transgenic - metabolism; Mutation; PLP; Pyridoxal Phosphate - metabolism; pyridoxine; Pyridoxine - pharmacology; Pyridoxine - therapeutic use; Recombinant Fusion Proteins - metabolism; Yeasts - genetics; Yeasts - metabolism
• ISSN: 1059-7794; 1098-1004
• DOI: 10.1002/humu.20320

Cystathionine β‐synthase (CBS) deficiency is a recessive genetic disorder characterized by extremely elevated levels in plasma homocysteine. Patients homozygous for the I278T or R266K mutations respond clinically to pharmacologic doses of pyridoxine, the precursor of a cofactor for the enzyme, 5′‐pyridoxal phosphate (PLP). Here we test the hypothesis that these mutations are pyridoxine responsive because they lower the affinity of the enzyme for PLP. We show that recombinant R266K has 30 to 100% of the specific activity of the wild‐type enzyme, while I278T only has only 1 to 5% activity. Kinetic studies show that the decreased activity in both enzymes is due to reduced turnover rate and not substrate binding. Neither I278T nor R266K appear to greatly affect multimer status of the enzyme. The R266K enzyme has reduced affinity for PLP compared to the wild‐type enzyme, providing a mechanism for the pyridoxine response observed in patients. Surprisingly, the I278T enzyme does not have altered affinity for PLP. To confirm that this was not an in vitro artifact, we examined pyridoxine response in mice that stably express human I278T as their sole source of CBS activity. These mice have extremely elevated plasma homocysteine levels and do not respond significantly to large doses of pyridoxine. Our findings suggest that there may be multiple mechanisms involved in response to pyridoxine. Hum Mutat 27(5), 474–482, 2006. © Published 2006 Wiley‐Liss, Inc.