Degradation of fluorene by Brevibacterium sp. strain DPO 1361: a novel C-C bond cleavage mechanism via 1,10-dihydro-1,10-dihydroxyfluoren-9-one.

Angular dioxygenation has been established as the crucial step in dibenzofuran degradation by Brevibacterium sp. strain DPO 1361 (V. Strubel, K. H. Engesser, P. Fischer, and H.-J. Knackmuss, J. Bacteriol. 173:1932-1937, 1991). The same strain utilizes biphenyl and fluorene as sole sources of carbon and energy. The fluorene degradation sequence is proposed to be initiated by oxidation of the fluorene methylene group to 9-fluorenol. Cells grown on fluorene exhibit pronounced 9-fluorenol dehydrogenase activity. Angular dioxygenation of the 9-fluorenone thus formed yields 1,10-dihydro-1,10-dihydroxyfluoren-9-one (DDF). A mechanistic model is presented for the subsequent C-C bond cleavage by an NAD(+)-dependent DDF dehydrogenase, acting on the angular dihydrodiol. This enzyme was purified and characterized as a tetramer of four identical 40-kDa subunits. The following Km values were determined: 13 microM for DDF and 65 microM for 2,3-dihydro-2,3-dihydroxybiphenyl. The enzyme also catalyzes the production of 3-(2'-carboxyphenyl)catechol, which was isolated, and structurally characterized, in the form of the corresponding lactone, 4-hydroxydibenzo-(b,d)-pyran-6-one. Stoichiometry analysis unequivocally demonstrates that angular dioxygenation constitutes the principal pathway in Brevibacterium sp. strain DPO 1361.

The Rhizobium meliloti rhizopine mos locus is a mosaic structure facilitating its symbiotic regulation.

The Rhizobium meliloti L5-30 mos locus, encoding biosynthesis of the rhizopine 3-O-methyl-scyllo-inosamine, is shown to be a mosaic structure. The mos locus consists of four open reading frames (ORFs) (ORF1 and mosABC) arranged in an operon structure. Within this locus, several domains of homology with other prokaryotic symbiotic genes (nifH, fixA, fixU, and nifT) are present, suggesting that this locus may represent a hot spot for rearrangement of symbiotic genes. Unusually, these domains are present in the coding as well as noncoding regions of the mos locus. Proteins corresponding to those encoded by mosABC, but not ORF1, have been detected in nodule extracts by using antibodies. As ORF1 shows extensive homology with the 5' region of the nifH gene (P.J. Murphy, N. Heycke, S.P. Trenz, P. Ratet, F.J. de Bruijn, and J. Schell, Proc. Natl. Acad. Sci. USA 85:9133-9137, 1988) and a frameshift mutation indicates that expression of this ORF is not required for mos activity, we propose that the mos locus has acquired a duplicated copy of nifH, including the promoter region, in order to become symbiotically regulated. Surprisingly, since the functions are likely different, MosA has an amino acid sequence similar to that of the DapA protein of Escherichia coli. The central domain of MosB has extensive homology with a range of diverse proteins involved with carbohydrate metabolism in either antibiotic or outer-cell-wall biosynthesis. This region is also common to the regulatory proteins DegT and DnrJ, suggesting a regulatory role for MosB. The structure of MosC is consistent with its being a membrane transport protein.

Synthesis of an opine-like compound, a rhizopine, in alfalfa nodules is symbiotically regulated.

We show that the promoter of the mos locus, which encodes genes required for the synthesis of a nodule-specific, opine-like compound, a rhizopine, in alfalfa nodules is regulated by the symbiotic nitrogen-fixation regulatory gene nifA. The 5'-regulatory region and amino-terminal end of the first open reading frame of the mos locus are highly homologous to the 5'-regulatory region and amino-terminal portion of the Rhizobium meliloti nifH gene. The coordinate regulation of mos and nif genes suggests that the mos locus plays a symbiotic role. We propose that the rhizopine enhances the survival of the bacterial partner in the symbiosis.