B12 METABOLISM IN HUMANS By NICOLE AURORA LEAL A ...

at the 5’-C of ATP by cob(I)alamin resulting in adenosylation (Fonseca et al. 2002).
Additionally, triphosphate was identified as a reaction byproduct of adenosylation and
was also found to be a strong inhibitor of the reaction (Fonseca et al. 2002).
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The archaeal T. acidophilum PduO-type ATR has been purified, crystallized, and
partially characterized (Saridakis et al. 2004). T. acidophilum ATR has 27% amino acid
identity to PduO. This ATR is specific for ATP and deoxy-ATP. Additionally, the
crystal structure revealed a trimeric configuration. Each of the subunits is composed of
five alpha helical domains that aid in subunit assembly due to hydrophobic, ionic, and
polar interactions depending on the domain. In contrast to CobA, there is no
recognizable P-loop motif suggesting a different binding method for ATP.
Formation of methylcobalamin
The synthesis CH3Cbl is coupled to cob(II)alamin reduction (Huennekens et al.
1982). Once cob(II)alamin is reduced, methylation is dependent on SAM as the methyl
donor for the formation of CH3Cbl. The E. coli cob(II)alamin reductase requires FldA
for the reductive activation of MS (Fujii and Huennekens 1974). Humans lack FldA,
requiring an alternate reductase for CH3Cbl formation.
Because of the role of FldA in bacteria, it was proposed that the human reductase
would contain binding sites for FMN and FAD (Leclerc et al. 1998). Using homology
based PCR, a cDNA with consensus sequences to predicted binding sites for FMN, FAD,
and NADPH was cloned and proposed to encode methionine synthase reductase (MSR),
the cob(II)alamin reductase required for MS activation (Leclerc et al. 1998). The MSR
gene (MTRR) was mapped to chromosome 5 at position p15.2 (Leclerc et al. 1999).
Analysis of the gene revealed that the N-terminal sequence is consistent with cytosolic
targeting, but alternative splicing at the 5’ end of MTRR mRNA generates a