Human Molecular Genetics, 2001, Vol. 10, No. 6 635-643
© 2001 Oxford University Press
Mutations in the regulatory domain of cystathionine ß–synthase can functionally suppress patient-derived mutations in cis
1Division of Population Science and 2Division of Basic Science, Fox Chase Cancer Center, Philadelphia, PA 19111, USA
Human cystathionine ß–synthase (CBS) is an S-adenosylmethionine-regulated enzyme that plays a key role in the metabolism of homocysteine. Mutations in CBS are known to cause homocystinuria, an inborn error in metabolism. We previously developed a yeast functional assay for CBS and used it to characterize mutations found in homocystinuric patients. We discovered that many patient-derived mutations are functionally suppressed by deletion of the C-terminal 142 amino acids, which contain a 53 amino acid motif known as the CBS domain. This domain is found in a wide variety of proteins of diverse biological function. Here we have used a genetic screen to identify missense mutations in the C-terminal region of CBS that can suppress the most common patient mutation, I278T. Seven suppressor mutations were identified, four of which map to the CBS domain. When combined in cis with another pathogenic mutation, V168M, six of seven of the suppressor mutations rescued the yeast phenotype. Enzyme activity analyses indicate that the suppressors restore activity from <2% to 17–64% of the wild-type levels. Analysis of the suppressor mutations in the absence of the pathogenic mutation shows that six of the seven suppressor alleles have lost enzymatic responsiveness to S-adenosylmethionine. Using homology modeling, we show that the suppressor mutations appear to map on one face of the CBS domain. Our results indicate that subtle changes to the C-terminus of CBS can restore activity to mutant proteins and provide a rationale for screening for compounds that can activate mutant CBS alleles.
+ To whom correspondence should be addressed at: Fox Chase Cancer Center, P3016, 7701 Burholme Avenue, Philadelphia, PA 19111, USA. Tel: +1 215 728 3030; Fax: +1 215 214 1623; Email: wd_kruger@fccc.edu
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