The opgGIH and opgC genes of Rhodobacter sphaeroides form an operon that controls backbone synthesis and succinylation of osmoregulated periplasmic glucans

Abstract

Osmoregulated periplasmic glucans (OPGs) of Rhodobacter sphaeroides are anionic cyclic molecules that accumulate in large amounts in the periplasmic space in response to low osmolarity of the medium. Their anionic character is provided by the substitution of the glucosidic backbone by succinyl residues. A wild-type strain was subject to transposon mutagenesis, and putative mutant clones were screened for changes in OPGs by thin layer chromatography. One mutant deficient in succinyl substitution of the OPGs was obtained and the gene inactivated in this mutant was characterized and named opgC. opgC is located downstream of three ORFs, opgGIH, two of which are similar to the Escherichia coli operon, mdoGH, governing OPG backbone synthesis. Inactivation of opgG, opgI or opgH abolished OPG production and complementation analysis indicated that the three genes are necessary for backbone synthesis. In contrast, inactivation of a gene similar to ndvB, encoding the OPG-glucosyl transferase in Sinorhizobium meliloti, had no consequence on OPG synthesis in Rhodobacter sphaeroides. Cassette insertions in opgH had a polar effect on glucan substitution, indicating that opgC is in the same transcription unit. Expression of opgIHC in E. coli mdoB/mdoC and mdoH mutants allowed the production of slightly anionic and abnormally long linear glucans.

The Brucella abortus Cyclic β-1,2-Glucan Virulence Factor Is Substituted with O-Ester-Linked Succinyl Residues

“…A gene with high homology to the opgC gene of Rhodobacter sphaeroides was identified in the B. abortus 2308 genome (12,14). The gene is highly conserved in B. melitensis and B. suis (19,34).…”

“…The opgC gene of R. sphaeroides and the mdoC gene of E. coli encode glucan succinyltransferases (14,29). Cgm, OpgC, and MdoC have similar sizes (393, 399, and 385 amino acids, respectively) and exhibit stretches of hydrophobic amino acids over their entire lengths.…”

“…(5Ј-T CATTTTCGGTGCGTGTTTT-3Ј) and cgm-rev. (5Ј-CATAAAAAAAGAGCC GCAAG-3Ј), designed according to the sequence of a B. abortus gene highly homologous to the R. sphaeroides OpgC gene (12,14).…”

“…Succinyl transfer was also shown to occur in the periplasmic compartment in S. meliloti (6). In E. coli and R. sphaeroides, the genes mdoC and opgC, respectively, encode predicted membrane proteins required for wild-type succinyl modification of OPG (14,29). E. coli has two phosphoglycerol transferases of glucans (I and II), one membrane bound and the second periplasmic.…”

Periplasmic glucans isolated from Proteobacteria

“…In the case of B. abortus, an open reading frame (BAB1_1781) homologous to R. sphaeroides glucans succinyltransferase (opgC) has been recently identified as the cgm gene, which encodes the enzyme responsible for succinyl modification of the PGs (28). The opgGIH and opgC genes in R. sphaeroides have been shown to form an operon that controls the backbone synthesis and succinylation of the PGs, respectively (45). A genetic locus, opgHXcv, in X. campestris pv.…”