Analysis of nucleotide methylation in DNA from Corynebacterium glutamicum and related species

Plasmid DNA (pCSL17) isolated from Corynebacterium glutamicum transformed recipient McrBC+ strains of Escherichia coli with lower efficiency than McrBC- strains, confirming a previous report by Tauch et al. (FEMS Microbiol. Lett. 123 (1994) 343-348) which inferred that C. glutamicum DNA contains methylcytidine. Analysis of nucleotides in C. glutamicum-derived chromosomal and plasmid DNA failed to detect significant levels of methylated adenosine, but methylated cytidine was readily detected. Restriction enzymes which are inhibited by the presence of methylcytidine in their recognition sequence failed to cut pCSL17 from C. glutamicum, whereas enzymes which require methylation at adenosine in GATC sequences failed to cut. Failure of HaeIII to cut two specific sites of C. glutamicum-derived pCSL17 identified the first cytidine in the sequence GGCCGC as one target of methylation in this species, which contains the methyltransferase recognition sequence. Although Brevibacterium lactofermentum-derived DNA showed a similar methylation pattern by HPLC analysis, HaeIII cleaved these GGCCGC sites, suggesting differences in the specificity of methylation between these two species. Results for all analyses of B. flavum DNA were identical to those for C. glutamicum.