Both the cobalamin-dependent and cobalamin-independent forms of the enzyme carry out the same overall chemical reaction, the transfer of a methyl group from 5-methyltetrahydrofolate (N5-MeTHF) to homocysteine, yielding tetrahydrofolate (THF) and methionine.

In humans, the enzyme is reduced in this process by methionine synthase reductase (MTRR), which consists of flavodoxin-like and ferrodoxin-NADP+ oxidoreductase (FNR)-like domains.

[20] The mechanism of the cobalamin-independent (MetE) form, by contrast, proceeds through a direct methyl transfer from the activated N5-MeTHF to zinc thiolate homocysteine.

The two domains adopt a face-to-face double barrel architecture, which requires a "closing" of the structure upon binding of both substrates to enable the direct methyl transfer.

[22] Substrate-binding strategies are similar to MetH, although in the case of MetE the zinc atom is instead coordinated to two cysteines, a histidine and a glutamate,[23] for which an example is shown on the right.

[29][30] As such, methionine synthase serves an essential function by allowing the SAM cycle to perpetuate without a constant influx of Met.

[31][32] In bacteria and plants, methionine synthase serves a dual purpose of both perpetuating the SAM cycle and catalyzing the final synthetic step in the de novo synthesis of Met, which is one of the 20 canonical amino acids.

[33][11] While the chemical reaction is exactly the same for both processes, the overall function is distinct from methionine synthase in humans because Met is an essential amino acid that is not synthesized de novo in the body.

Most cases of methionine synthase deficiency are symptomatic within 2 years of birth with many patients rapidly developing severe encephalopathy.

[37] One consequence of reduced methionine synthase activity that is measurable by routine clinical blood tests is megaloblastic anemia.