Defects in human methionine synthase in cblG patients

Inborn errors resulting in isolated functional methionine synthase deficiency fall into two complementation groups, cblG and cblE. Using biochemical approaches we demonstrate that one cblG patient has greatly reduced levels of methionine synthase while in another, the enzyme is specifically impaired in the reductive activation cycle, The biochemical data suggested that low levels of methionine synthase activity in the first patient may result from mutations in the catalytic domains of the enzyme, reduced transcription, or generation of unstable message or protein, Using Northern analysis, we demonstrate that the molecular basis for the biochemical phenotype in this patient is associated with greatly diminished steady-state levels of methionine synthase mRNA, The biochemical data on the second patient cell line implicated mutations specific to reductive activation, a function that is housed in the C-terminal AdoMet-binding domain and the intermediate B-12-binding domain, in the highly homologous bacterial enzyme, We have detected two mutations in a compound heterozygous state, one that results in conversion of a conserved proline (1173) to a leucine residue and the other a deletion of an isoleucine residue (881), The crystal structure of the C-terminal domain of the Escherichia coli MS predicts that the Pro to Leu mutation could disrupt activation since it is embedded in a sequence that makes direct contacts with the bound AdoMet, Deletion of isoleucine in the B-12-binding domain would result in shortening of a beta-sheet, Our data provide the first evidence for mutations in the methionine synthase gene being culpable for the cblG phenotype. In addition, they suggest directly that mutations in methionine synthase can lead to elevated homocysteine, implicated both in neural tube defects and in cardiovascular diseases.