title: Cystathionine beta-synthase mutations: effect of mutation topology on folding and activity.
abstract: Misfolding of mutant enzymes may play an important role in the pathogenesis of cystathionine beta-synthase (CBS) deficiency. We examined properties of a series of 27 mutant variants, which together represent 70% of known alleles observed in patients with homocystinuria due to CBS deficiency. The median amount of SDS-soluble mutant CBS polypeptides in the pellet after centrifugation of bacterial extracts was increased by 50% compared to the wild type. Moreover, mutants formed on average only 12% of tetramers and their median activity reached only 3% of the wild-type enzyme. In contrast to the wild-type CBS about half of mutants were not activated by S-adenosylmethionine. Expression at 18 degrees C substantially increased the activity of five mutants in parallel with increasing the amounts of tetramers. We further analyzed the role of solvent accessibility of mutants as a determinant of their folding and activity. Buried mutations formed on average less tetramers and exhibited 23 times lower activity than the solvent exposed mutations. In summary, our results show that topology of mutations predicts in part the behavior of mutant CBS, and that misfolding may be an important and frequent pathogenic mechanism in CBS deficiency.
authors.affiliation: First Faculty of Medicine, Charles University in Prague and General University Hospital in Prague, Institute of Inherited Metabolic Disorders, Prague, Czech Republic. viktor.kozich@lf1.cuni.cz
authors.forename: Viktor
authors.initials: V
authors.lastname: Kozich
authors.forename: Jitka
authors.initials: J
authors.lastname: Sokolová
authors.forename: Veronika
authors.initials: V
authors.lastname: Klatovská
authors.forename: Jakub
authors.initials: J
authors.lastname: Krijt
authors.forename: Miroslav
authors.initials: M
authors.lastname: Janosík
authors.forename: Karel
authors.initials: K
authors.lastname: Jelínek
authors.forename: Jan P
authors.initials: JP
authors.lastname: Kraus
mesh_terms: D015153:Blotting, Western; D002384:Catalysis; D020134:Catalytic Domain / Q000235:genetics; D003080:Cold Temperature; D003541:Cystathionine beta-Synthase / Q000737:chemistry / Q000172:deficiency / Q000235:genetics*; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D006712:Homocystinuria / Q000201:enzymology / Q000235:genetics*; D006801:Humans; D008958:Models, Molecular; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D017510:Protein Folding; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D012995:Solubility
chemical_list: D050505:Mutant Proteins; D003541:Cystathionine beta-Synthase