DNA sequence determination and characterization of plasmid pRC12 isolated from Lactobacillus curvatus CRL 705

TERÁN, L.; ARISTIMUÑO, C.; FADDA, S.; SESMA, F.; HEBERT, E.M.; RAYA, R.R.

San M de Tucumán, Tucumán, Argentina.

Simposio; IV International Symposium on Lactic Acid Bacteria: Food, Health and Applications.; 2013

Lactobacillus curvatus CRL 705 has a potential great importance in food industry. This strain which was isolated from a traditional Argentinian sausage harbors two plasmids: pRC18 and pRC12. Plasmid pRC18 was associated with the production of the bacteriocins Lac705 and AL 705 (with antilisteria activity), while plasmid pRC12 was proposed to be associated with sucrose metabolism.Here, the genome DNA sequence of plasmid pRC12 was determined and annotated according to standard operating procedures (SOPs) for prokaryotic annotation from ISGA, the RAST annotation server, Glimmer 3.02 modeling software package, and tRNAscan-SE 1.21. Finally, the DNA sequence was compared to those present in the GenBank database using BLAST. The DNA sequence of plasmid pRC12 consists of 12342 bp with a mol% G + C content of 44%. Thirteen putative protein encoding open reading frames (orf) of more than 70 nucleotides in length were identified. Three encoding putative proteins involved in the replication process were identified: Orf4, Orf6 and Orf7 with homologies to the initiation protein RepA of Lactobacillus coryniformis subsp. torquens KCTC 3535 (identity 98%), replication protein of Pediococcus acidilactici (identity 98%), and replication initiation protein RepA of Lactobacillus pentosus IG1(identity 99%), respectively. It also has three orf related to DNA mobilization: orf3 which encodes for a protein highly similar to the relaxase/mobilization domain of Lactobacillus pentosus IG1, orf9 which encodes for a protein with homology to MobC of Lactobacillus hilgardii ATCC 8290 and the orf13 that encodes for a protein highly similar to MobC of Lactobacillus buchneri ATCC 11577. Furthermore, orf10 and orf11, twogenes organized in the same orientation, encode proteins for the toxin of the toxin-antitoxin stabilibty system of Pediococcus claussenii ATCC BAA-344 and for the antitoxin of the toxin-antitoxin system of Lactobacillus vini DSM 20605, respectively. The protein encode by orf8 has homology to the toxin component of the toxin-antitoxin system of the RelE family of the Enterococus faecium R497. The deduced amino acid product of orf5 showed the highest identity to the integrase of Lactobacillus coryniformis subsp. torquens KCTC 3535 and orf1 encodes for the putative protein of calcium-transporting ATPase of Lactobacillus casei W56. The Orf2 protein was detected only with distant relationship with hydrolase of Enterococcus faecium E1071, which we proposed to be associated with the sucrose metabolism. Finally, orf12 encodes one additional putative protein of unknown function. Functional characterizations of plasmid pRC12 are being carried out; these studies will allow us to expand the biotechnological applications of strain L. curvatus CRL 705.