Extreme Halophiles Synthesize Betaine from Glycine by Methylation

Abstract

Glycine betaine is a compatible solute, which is able to restore and maintain osmotic balance of living cells. It is synthesized and accumulated in response to abiotic stress. Betaine acts also as a methyl group donor and has a number of important applications including its use as a feed additive. The known biosynthetic pathways of betaine are universal and very well characterized. A number of enzymes catalyzing the two-step oxidation of choline to betaine have been isolated. In this work we have studied a novel betaine biosynthetic pathway in two phylogenically distant extreme halophiles, Actinopolyspora halophila and Ectothiorhodospira halochloris. We have identified a three-step series of methylation reactions from glycine to betaine, which is catalyzed by two methyltransferases, glycine sarcosine methyltransferase and sarcosine dimethylglycine methyltransferase, with partially overlapping substrate specificity. The methyltransferases from the two organisms show high sequence homology. E. halochloris methyltransferase genes were successfully expressed in Escherichia coli, and betaine accumulation and improved salt tolerance were demonstrated.

Isolation and Functional Characterization ofN-Methyltransferases That Catalyze Betaine Synthesis from Glycine in a Halotolerant Photosynthetic Organism Aphanothece halophytica

“…Thus, the biogenesis of Ntrimethyl compounds occurs via diversity methyltransferases. Despite of diversity methyltransferases, betaine synthesis from glycine was only observed in halotolerant organisms (15). This suggests that the methyltransferase genes, encoding ApGSMT and ApDMT, in A. halophytica might evolve independently from those of methyltransferases in choline biogenesis and mammalian glycine N-methyltransferase.…”

“…Homologous sequences to ApGSMT and ApDMT were only found in E. halochloris and A. halophila. A. halophytica methyltransferases were different from that of A. halophila in which one gene was found (15). ApGSMT and ApDMT of A. halophytica were similar to EcGSMT and EcSDMT of E. halochloris, respectively, but substrate specificity of ApDMT was different from EcSDMT (Fig.…”

“…The mutated amino acid, Arg (R), was also marked above the alignment. AcGSDMT, A. halophila methyltransferase (15). B, alignment of the deduced amino acid sequences of ORF2.…”

“…Betaine synthesis from simple carbon sources has also been suggested in aerobic heterotrophic eubacterium Actinopolyspora halophila (13) and halotolerant cyanobacterium of Aphanothece halophytica (14). Recently, the methyltransferase genes that are involved in betaine synthesis have been isolated from E. halochloris and A. halophila (15). Two methyltransferase genes were involved in E. halochloris.…”

“…One of gene products catalyzed the methylation reactions of glycine and sarcosine to sarcosine and dimethylglycine, respectively (EcGSMT), 1 whereas the other one catalyzed the methylations of sarcosine and dimethylglycine to dimethylglycine and betaine, respectively (EcSDMT) (15,16). By contrast, one ORF was found in A. halophila of which the N-and C-terminal parts had homologous sequences to those of EcGSMT and EcSDMT, respectively (15). The functionality of A. halophila methyltransferase was not well shown due to the formation of cell pellet when expressed in Escherichia coli.…”