REPLICATION REGIONS OF 2 PAIRS OF INCOMPATIBLE LACTOCOCCAL THETA-REPLICATING PLASMIDS

Incompatibility tests were performed employing 12 replicons belonging to a family of homologous lactococcal theta-replicating plasmids. Two pairs of incompatible plasmids were found namely, pFV1001 and pFV1201, and pJW565 and pFW094. The replicons of plasmids pFV1001, pFV1201, pJ W565, pJW566, and pFW094 were sequenced. Alignments were made of the r eplicational origins (repA) and putative replication proteins (RepB) o f these and ii related plasmid sequences. Comparison of the alignments with the incompatibility data indicated that the incompatibility dete rminant could be contained within the 22-bp tandem repeats DRII and/or the inverted repeat IR1 in repA, In support the incompatibility deter minant of pJW563 was localized to a 743-bp fragment encompassing repA. A stretch of 13 amino acids of RepB was proposed to be responsible fo r the plasmid-specific initiation of replication. This stretch is pan, of a domain containing features that are highly conserved within the proposed DNA binding regions of the initiation proteins from several w ell-characterized plasmids from Gram-negative bacteria, including pSC1 01, R6K, and mini-F. (C) 1997 Academic Press.