Análisis genómico del cluster biosintético de vitamina B12 (cobalamina) en Lactobacillus coryniformis CRL 1001

Abstract

The vitamin B12 (cobalamin-CBL), a very complex non-polymeric macromolecule synthesized only by some bacteria and archaea, and essential to humans and animals. The highly complicated CBL biosynthesis involves about 30 synthesis steps. We demonstrated that cell extract of Lactobacillus coryniformis CRL 1001 is able to correct the coenzyme B12 requirement of Salmonella enterica serovarTyphimuriumin minimal medium. The aim of this study was the sequencing of CRL1001 genome and the molecular characterization of CBL biosynthesis in this strain. L. coryniformis CRL 1001 genome was sequenced by a whole-genome shotgun (WGS) strategy with an Ion Torrent personal genome machine based upon libraries created using NEBNext DNA library kits. Genomic analysis was done using the RAST annotation Server, Blast algorithms, ISGA and KEGG databases.The draft genome sequence consists of 2,829,178 bases with a mean GC content of 42%. A total of 3,341 coding sequences (CDS) and 82 structural RNAs (58 tRNAs) were predicted. RAST analysis evidenced the presence of at least 30 genes (cob genes) involved in the CBL biosynthesis in CRL 1001 strain. This finding is the first evidence for cobalamin biosynthesis genes in this species. Comparative studies among vitamin B12 producer strains demonstrated that the genetic organization of cob operon is conserved in this strain and these genes are adjacent to the pdu operon and pocR gene. The hem genes (hem A, C, B y L) present in L. coryniformis CRL 1001 genome are located among cob operon in similar way to anaerobic microorganisms. Interestingly, the cblT y cblS genes were identified in CRL1001 strain genome. These genes encode a putative protein kinase and a α-ribasol transporter, respectively. The cblT y cblS genes are present in CBLproducers Listeria sp. strains but absent in CBL-producers L. reuteri strains. The knowledge of cob genes and their regulation in vitamin producer lactic strains constitutes an interesting biotechnological alternative for developing fortified foods.