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Cystathionine β-synthase (CBS) is a key enzyme in sulfur metabolism, and its inherited deficiency causes homocystinuria. Mammalian CBS is modulated by the binding of S-adenosyl-l-methionine (AdoMet) to its regulatory domain, which activates its catalytic domain. To investigate the underlying mechanism, we performed x-ray crystallography, mutagenesis, and mass spectrometry (MS) on human CBS. The 1.7 Å structure of a AdoMet-bound CBS regulatory domain shows one AdoMet molecule per monomer, at the interface between two constituent modules (CBS-1, CBS-2). AdoMet binding is accompanied by a reorientation between the two modules, relative to the AdoMet-free basal state, to form interactions with AdoMet via residues verified by mutagenesis to be important for AdoMet binding (Phe(443), Asp(444), Gln(445), and Asp(538)) and for AdoMet-driven inter-domain communication (Phe(443), Asp(538)). The observed structural change is further supported by ion mobility MS, showing that as-purified CBS exists in two conformational populations, which converged to one in the presence of AdoMet. We therefore propose that AdoMet-induced conformational change alters the interface and arrangement between the catalytic and regulatory domains within the CBS oligomer, thereby increasing the accessibility of the enzyme active site for catalysis.

Original publication

DOI

10.1074/jbc.M114.610782

Type

Journal article

Journal

J Biol Chem

Publication Date

26/12/2014

Volume

289

Pages

36018 - 36030

Keywords

Activation, Allosteric Regulation, Conformational Change, Crystallography, Cystathionine β-Synthase, Enzyme, Mass Spectrometry (MS), S-adenosyl-l-methionine (AdoMet), SAM, Catalytic Domain, Crystallography, X-Ray, Cystathionine beta-Synthase, Humans, Hydrogen Bonding, Models, Molecular, Protein Binding, Protein Structure, Secondary, S-Adenosylmethionine