GENOME STRUCTURE OF A LACTOBACILLUS PLASMID PNMO BELONGING TO THE ROLLING-CIRCLE TYPE REPLICON

The new small cryptic plasmid pNMO propagating in Lactobacillus casei IAM1045 was isolated from a naturally occurring Lactobacillus strain N 1f, Sequencing analysis revealed that the pNMO genome is a double-stra nded DNA of 3522 bp, and has two possible open reading frames, rep and pre as well as three other minor ORFs. The predicted rep product (Rep ) of 318 amino acids bears close resemblance to other presumed replica tion proteins (Rep) encoded by the Lactobacillus plasmids such as pLP1 , replicating by the rolling-circle (RC) mechanism via single-stranded (SS) DNA intermediates. Upstream rep, pNMO carries two putative repli cation origins: one is a Rep-mediated nick-site for the plus-strand sy nthesis (+ori), and the other is a single strand origin (sso) for the minus-strand replication. Preliminary in vivo experiments suggested th at pNMO generates SS DNA intermediates in the N1f cells. On the other hand, the putative pNMO pre product (Pre) of 361 amino acids shows ext ensive similarity to other presumed Pre/Mov (plasmid recombination/mob ilization) proteins encoded by the Lactobacillus plasmids pLB4 and pLA B1000. These results suggested that pNMO replicates via the RC mechani sm, and carries the Pre/Mov system.