No abstract

Methylation of a base in a specific DNA sequence protects the DNA from nucleolytic cleavage by restriction enzymes recognizing the same sequence. The MboII restriction-modification (R-M) system of Moraxella bovis ATCC 10900 consists of a restriction endonuclease gene and two methyltransferase genes. The enzymes encoded by this system recognize an asymmetrical sequence 59-GAAGA-39/39-CTTCT-59. M1.MboII modifies the last adenine in the recognition sequence 59-GAAGA-39 to N 6 -methyladenine. A second methylase, M2.MboII, was cloned and purified to electrophoretic homogeneity using a four-step chromatographic procedure. It was demonstrated that M2.MboII modifies the internal cytosine in the recognition sequence 39-CTTCT-59, yielding N 4 -methylcytosine, and moreover is able to methylate single-stranded DNA. ) inhibit M2.MboII methylation activity. It was found that the isomethylomer M2.NcuI from Neisseria cuniculi ATCC 14688 behaves in the same manner. Functional analysis showed that the complete MboII R-M system, consisting of two methyltransferases genes and the mboIIR gene, is the most stable and the least harmful to bacterial cells.

No abstract

We report the characterization of a small cryptic plasmid unlike any previously described from Moraxella bovis ATCC 10900, a Gram-negative bacterium belonging to the family Moraxellaceae. The complete nucleotide sequence of the plasmid pMbo4.6 was determined. The plasmid was analyzed and found to be 4658 in size with a G+C content of 38.6 mol %. Computer analysis of the sequence data revealed four major open reading frames encoding putative proteins of 10.1 (ORF1), 64.2 (ORF2), 45.7 (ORF3), and 12.1 kDa (ORF4). ORF1 and ORF2 encode proteins that show a high level of amino acid sequence similarity (44 %) with some mobilization proteins. ORF3 encodes a protein showing a relatively high amino acid sequence similarity (about 40 %) with several plasmid replication initiator proteins. Upstream of ORF3, a 320-bp intergenic region, constituting the putative origin of replication that contained an AT-rich region followed by four direct repeats, was identified. This set of repeated sequences resembles iteron structures and plays an important role in the control of plasmid replication by providing a target site for the initiation of transcription and replication factors (IHF and RepA). Several palindromic sequences, inverted repeats, and hairpin-loop structures, which might confer regulatory effects on the replication of the plasmid, were also noted. ORF4 encodes an uncharacterized protein, conserved in bacteria, belonging to the DUF497 family. Sequence analysis and structural features indicate that pMbo4.6 replicates by a theta mechanism.