Methionine γ-lyase Traps Cofactor Pyridoxal-5'-phosphate through Specific Serine-Mediated Affinity

Pyridoxal-5'-phosphate (PLP), the ubiquitous and ancient cofactor, plays important roles in enzymatic elimination, transamination and other reactions. The catalytic efficiency of PLP-dependent enzymes is significantly higher than that of free PLP. The recruitment of PLP from the environment by the enzymes, particularly through interactions outside the active site, is the key step determining the occurrence of PLP-mediated catalysis. However, the precise mechanism by which enzymes recruit PLP remains elusive. Methionine {gamma}-lyase (MGL), a PLP-dependent enzyme, catalyzes the degradation of L-methionine, thereby suppressing cancer cell proliferation through serum or dietary methionine depletion. Here, we report the crystal structure of yMGL, which belongs to a newly identified subgroup of cystathionine {gamma}-lyases, in complex with L-methionine and PLP. Through truncating the C-terminal domain of yMGL both in vitro and in vivo, we demonstrated that this domain, outside the canonical PLP-binding domain, is essential for the specific interaction between yMGL and PLP, as well as for efficient L-methionine catabolism. A conserved Ser in the C-terminal domain, positioned at the active-site entrance of MGLs, confers specificity for PLP. These findings elucidate a previously uncharacterized mechanism of PLP recruitment by MGLs and promote rational design of MGLs with tailored cofactor selectivity and catalytic performance.