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The methylotrophic proteobacterium Methylobacterium extorquens AM1 possesses tetrahydromethanopterin (H₄MPT)-dependent enzymes, which are otherwise specific to methanogenic and sulfate-reducing archaea and which have been suggested to be involved in formaldehyde oxidation to CO₂ in M. extorquens AM1. The distribution of H₄MPT-dependent enzyme activities in cell extracts of methylotrophic bacteria from 13 different genera are reported. H₄MPT-dependent activities were detected in all of the methylotrophic and methanotrophic proteobacteria tested that assimilate formaldehyde by the serine or ribulose monophosphate pathway. H₄MPT-dependent activities were also found in autotrophic Xanthobacter strains. However, no H₄MPT-dependent enzyme activities could be detected in other autotrophic α-proteobacteria or in gram-positive methylotrophic bacteria. Genes encoding methenyl H₄MPT …