Characterization and replication properties of the Zymomonas mobilis ATCC 10988 plasmids pZMO1 and pZM02

The complete nucleotide sequences of two small cryptic Zymomonas mobilis AT CC 10988 plasmids (pZMO1 and pZMO2) were determined. The plasmids showed 67 % homology to each other at their nucleotide level. Plasmid pZMO1 was 1651 bp long with 38% G + C content and contained an open reading frame (ORFZMO1 ) of 1044 nucleotides. ORFZMO1 is predicted to encode a polypeptide of 348 amino acids and shows a high degree of homology with gram-negative replicat ion proteins of rolling circle replicating plasmids, which belong to the pC 194/pUB110 family. Plasmid pZMO2 was found to be 1669 bp long, with a 38.5% G + C content, and it contained an ORF of 552 nucleotides (ORFZMO2) encodi ng a putative polypeptide of 184 amino acids. This polypeptide also shows a high degree of homology with the replication proteins of RCR plasmids of g ram-negative bacteria, but only at their N-termini. The region necessary fo r replication of both plasmids was determined by stability tests under nons elective conditions, following cloning in pBR325 and introduction in Z. mob ilis ATCC 10988 by pRK2013 assisted conjugation. Double- and single-strand origin regions were predicted by sequence analysis. Detection of single-str anded DNA in the extract of exponentially growing cells confirmed experimen tally the rolling circle replication mode of at least pZMO2. (C) 2000 Acade mic Press.