Recognition of two intracellular cobalamin binding proteins and their identification as methylmalonyl-CoA mutase and methionine synthetase.

Abstract

The granulocyte R-type cobalamin binding protein delivers cobalamin (Cbl)

MeaB Is a Component of the Methylmalonyl-CoA Mutase Complex Required for Protection of the Enzyme from Inactivation

“…Enzyme Assays-Enzyme activities were determined in M. extorquens AM1 and B. fungorum LB400 crude extracts obtained by passing cells through a French pressure cell at 1.2 ϫ 10 8 Pa, followed by centrifugation for 20 min at ϳ15,000 ϫ g. Methylmalonyl-CoA mutase activity was determined by two methods: a radiolabel-based assay (35) was used to monitor the MCM activity in the mutants, and an HPLC method (36) was used to characterize the MCM activity of purified proteins. To test for the presence of AdoCbl in cell extracts of the mutants and the wild type, 0.1 M apoMCM was added to the reaction mixture, and MCM activity was monitored by the radiolabeling assay.…”

Protection of aquo/hydroxocobalamin from reduced glutathione by a B12 trafficking chaperone

“…The sixth upper axial ligand determines the biological activities of B12 derivatives. In human cells, adenosylcobalamin (AdoCbl) and methylcobalamin (MeCbl), which are the active forms of B12, are used as the enzyme cofactors of methylmalonyl-CoA mutase in mitochondria and methionine synthase in cytosol, respectively (4,5). Hence, any form of B12 introduced into cells must be converted to AdoCbl and MeCbl via intracellular B12 metabolism.…”

Human ATP:Cob(I)alamin Adenosyltransferase and Its Interaction with Methionine Synthase Reductase

“…Humans are incapable of synthesizing AdoCbl and CH 3 Cbl de novo and depend on dietary sources of complex cobalamin precursors, such as vitamin B 12 (cyanocobalamin) and hydroxycobalamin (HOCbl). The pathway by which these precursors are metabolized to the coenzyme forms is thought to be similar in both prokaryotes and eukaryotes (Fig.…”

“…1) (5). The second B 12 dependent enzyme known in humans is CH 3 Cbl-dependent methionine synthase (MS). This enzyme is found in the cytoplasm where it catalyzes the conversion of methyltetrahydrofolate and homocysteine to tetrahydrofolate and methionine (6).…”

“…1) (8 -13). For the synthesis of CH 3 Cbl, cob(II)alamin associated with MS is reductively methylated to form CH 3 Cbl (6,14,15). For this reaction (which is also used for the reductive activation of MS following adventitious oxidation of the cobalamin cofactor), S-adenosylmethionine is the methyl-group donor and methionine synthase reductase (MSR) reduces cob(II)alamin to cob(I)alamin (16,17).…”