Domain Architecture of the Heme-Independent Yeast Cystathionine β-Synthase Provides Insights into Mechanisms of Catalysis and Regulation

Cystathionine β-synthase from yeast (Saccharomyces cerevisiae) provides a model system for understanding some of the effects of disease-causing mutations in the human enzyme. The mutations, which lead to accumulation of l-homocysteine, are linked to homocystinuria and cardiovascular diseases. Here we characterize the domain architecture of the heme-independent yeast cystathionine β-synthase. Our finding that the homogeneous recombinant truncated enzyme (residues 1−353) is catalytically active and binds pyridoxal phosphate stoichiometrically establishes that the N-terminal residues 1−353 compose a catalytic domain. Removal of the C-terminal residues 354−507 increases the specific activity and alters the steady-state kinetic parameters including the K d for pyridoxal phosphate, suggesting that the C-terminal residues 354−507 compose a regulatory domain. The yeast enzyme, unlike the human enzyme, is not activated by S-adenosyl-l-methionine. The truncated yeast enzyme is a dimer, whereas the full-length enzyme is a mixture of tetramer and octamer, suggesting that the C-terminal domain plays a role in the interaction of the subunits to form higher oligomeric structures. The N-terminal catalytic domain is more stable and less prone to aggregate than full-length enzyme and is thus potentially more suitable for structure determination by X-ray crystallography. Comparisons of the yeast and human enzymes reveal significant differences in catalytic and regulatory properties.