Molecular characterization of the bet genes encoding glycine betaine synthesis in Sinorhizobium meliloti 102F34

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Abstract
As a first step towards the elucidation of the molecular mechanisms responsible for the utilization of choline and glycine betaine (betaine) either as carbon and nitrogen sources or as osmoprotectants inSinorhizobium meliloti,we selected a Tn5 mutant, LTS23-1020, which failed to grow on choline but grew on betaine. The mutant was deficient in choline dehydrogenase (CDH) activity, failed to oxidize [methyl-14C]choline to [methyl-14C]betaine, and did not use choline, but still used betaine, as an osmoprotectant. The Tn5 mutation in LTS23-1020 was complemented by plasmid pCH034, isolated from a genomic bank ofS. meliloti102F34. Subcloning and DNA sequencing showed that pCH034 harbours two ORFs which showed 60% and 57% identity with theEscherichia coli betBgene encoding betaine-aldehyde dehydrogenase (BADH) andbetAgene encoding CDH, respectively. In addition to the homology withE. coligenes, the deduced sequence of the sinorhizobial BADH protein displays consensus sequences also found in plant BADHs. The deduced sequence of the sinorhizobial CDH protein shares only 21% identical residues with choline oxidase fromArthrobacter globiformis.The structural organization of thebetBAgenes inS. melilotidiffers from that described inE. coli: (i) the two ORFs are separated by a 210 bp sequence containing inverted repeats ressembling a putative rho-independent transcription terminator, and (ii) no sequence homologous tobetT(high-affinity choline transport system) orbetI(regulator) was found in the vicinity of the sinorhizobialbetBAgenes. Evidence is also presented that theS. meliloti betBAgenes are not located on the megaplasmids.