Georgia Edmonds
Research Mentor: Ruma Banerjee
Mentor Department: Biological Chemistry, Medicine
Author(s): Georgia Edmonds, Markus Ruetz, Ruma Banerjee
Session: Session 2 (10:00 AM – 10:50 AM)
Presentation Type: Poster 50
Abstract
ATP:cob(I)alamin adenosyltransferase (also known as MMAB) is a mitochondrial enzyme that synthesizes 5′-deoxyadenosylcobalamin (AdoCbl), the active cofactor form of vitamin B12 required by methylmalonyl-CoA mutase (MMUT). MMUT is essential for breaking down odd-chain fatty acids, cholesterol, and branched-chain amino acids. In addition to catalyzing AdoCbl formation, MMAB functions as a chaperone that delivers the cofactor to MMUT and repairs inactive enzyme. Mutations in MMAB disrupt this process and lead to methylmalonic aciduria type B, a metabolic disorder characterized by toxic accumulation of methylmalonic and propionic acids. In this study, we investigated how two patient-associated MMAB mutations, A135T and C132R, located in a loop outside the active site, alter cofactor binding, catalysis, and transfer. Spectrophotometric binding assays showed that the A135T mutation weakens AdoCbl affinity relative to wild-type MMAB by 4-fold, whereas the C132R variant binds with AdoCbl comparable affinity. In contrast, A135T MMAB accelerated Co–C bond cleavage and aquocobalamin (OH2Cbl) formation, while C132R MMAB displayed near-wild-type kinetics. AdoCbl transfer assays revealed that A135T MMAB transfers cofactor to MMUT more rapidly, with biphasic kinetics indicating both direct transfer and premature release into solution. C132R MMAB exhibited transfer rates similar to wild-type MMAB. Regulation by the GTPase MMAA was preserved for both mutants with GDP supporting transfer and GMPPCP (a non-hydrolyzable analog) blocking it. Finally, NADPH-dependent AdoCbl synthesis showed that both variants retain catalytic activity within the wild-type range. Together, these results reveal that A135T MMAB destabilizes the AdoCbl-bound state, promoting premature cofactor release and impairs delivery to MMUT. In contrast, C132R MMAB maintains near-normal function, suggesting that protein stability might be affected.
