The giant linear plasmid pHG207 from Rhodococcus sp. encoding hydrogen autotrophy: characterization of the plasmid and its termini

Abstract

As described previously, in Rhodococcus sp. (formerly Nocardia opaca) strains MRll and MR22, the ability to grow as an aerobic hydrogen bacterium (the Aut character) is located on giant conjugative linear plasmids -on pHG201 (270 kb) in strain MRll and on pHG205 (280 kb) in strain MR22. In an autotrophic transconjugant originating from MR22 a smaller plasmid, pHG207 (225 kb), was detected and shown to be a recombination product of the wild-type plasmids pHG204 and pHG205. A donor carrying pHG207 as the sole plasmid transferred the Aut marker at a 1000-fold frequency compared to the wild-type plasmid pHG205. Analysis of the plasmid ends revealed that plasmid pHG207 carries proteins at both ends; the proteins are linked to the 5' ends of the strands. The cloned end fragments of about 2 kb were sequenced and found to contain highly homologous sequences within the terminal 583 bp (left end part) and 560 bp (right end part). Several potential reading frames were detected, but database searching gave no indication about possibIe functions.

TraA is required for megaplasmid conjugation in Rhodococcus erythropolis AN12

“…Several diVerent Rhodococcus megaplasmids are known to be conjugative (Dabrock et al, 1994;Desomer et al, 1988;Kalkus et al, 1993;Priefert et al, 2004;Shimizu et al, 2001). The sequences and gene annotations of Wve rhodococcal megaplasmidsp33701 from Rhodococcus equi ATCC33701 (Takai et al, 2000), pBD2 from Rhodococcus erythropolis BD2 (Stecker et al, 2003), pRHL3 from Rhodococcus sp.…”

A linear megaplasmid, p1CP, carrying the genes for chlorocatechol catabolism of Rhodococcus opacus 1CP

“…Even plasmids with very similar ends may be the product of a recombination process, as was shown for pHG207. The two ends of pHG207 share 95 and 91 % identical base pairs over two regions of 237 and 329 positions, respectively; however, restriction analysis confirmed that the left and right end in fact originate from pHG204 and pHG205, respectively (Kalkus et al, 1993).…”

“…The R. opacus wild-type strains MR11 and MR22 were each shown to contain three linear plasmids, pHG201 (270 kb), pHG202 (400 kb) and pHG203 (420 kb), as well as pHG204 (190 kb), pHG205 (280 kb) and pHG206 (500 kb), respectively (Kalkus et al, 1990). Two of these plasmids, pHG201 and pHG205, as well as pHG207, obtained from the transconjugant strain MR2253 (Kalkus et al, 1993), enable their hosts to grow chemolithoautotrophically on gaseous hydrogen and carbon dioxide. Furthermore, pHG204 correlates with thallium resistance.…”

“…The fact that only one of the two ends of plasmids pHG201, pRHL2 and pBD2 is detected by BLASTN searches reflects the fact that linear plasmids of rhodococci, like their counterparts from streptomycetes (Chen et al, , 1996Lin et al, 1993;Pandza et al, 1998), may exchange their ends (Kalkus et al, 1993). Even plasmids with very similar ends may be the product of a recombination process, as was shown for pHG207.…”

“…We tested the electrophoretic immobility after omission of proteolytic and denaturating treatments in plasmid preparation or separation (Picardeau & Vincent, 1998) as well as the protection of protein-bearing DNA against exonuclease digestion (Kalkus et al, 1993).…”