Mapping peptides correlated with transmission of intrasteric inhibition and allosteric activation in human cystathionine beta-synthase.
Cystathionine beta-synthase plays a key role in the intracellular disposal of homocysteine and is the single most common locus of mutations associated with homocystinuria. Elevated levels of homocysteine are correlated with heart disease, Alzheimer's and Parkinson's diseases, and neural tube defects. Cystathionine beta-synthase is modular and subjected to complex regulation, but insights into the structural basis of this regulation are lacking. We have employed hydrogen exchange mass spectrometry to map peptides whose motions are correlated with transmission of intrasteric inhibition and allosteric activation. The mass spectrometric data provide an excellent correlation between kinetically and conformationally distinguishable states of the enzyme. We also demonstrate that a pathogenic regulatory domain mutant, D444N, is conformationally locked in one of two states sampled by the wild type enzyme. Our hydrogen exchange data identify surfaces that are potentially involved in the juxtaposition of the regulatory and catalytic domains and form the basis of a docked structural model for the full-length enzyme.
Sen
Suvajit
S
Biochemistry Department, University of Nebraska, Lincoln, Nebraska 68588-0664, USA.
Yu
Jiong
J
Yamanishi
Mamoru
M
Schellhorn
Daniel
D
Banerjee
Ruma
R
D000494:Allosteric Regulation / Q000235:genetics*; D000595:Amino Acid Sequence; D003541:Cystathionine beta-Synthase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004789:Enzyme Activation; D015971:Gene Expression Regulation, Enzymologic; D006710:Homocysteine / Q000378:metabolism; D006712:Homocystinuria / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D013058:Mass Spectrometry / Q000379:methods; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D010449:Peptide Mapping; D017434:Protein Structure, Tertiary
D006710:Homocysteine; D003541:Cystathionine beta-Synthase