G Posfai | PubMed Health

PubMed1986

Overproduction of the Bacillus sphaericus R modification methylase in Escherichia coli and its purification to homogeneity.

A DNA fragment containing the information coding for the GGCC-specific Bacillus sphaericus R modification methylase, BspR, was inserted into plasmid vector pKK223-3 under the control of the strong and inducible tac promoter, and transformed into Escherichia coli HB101. Upon induction this strain accumulated the methylase enzyme (while cell growth was inhibited) up to several percent of total cellular protein. Homogeneous methylase could be prepared in three purification steps.

Bacillus↗

PubMed1985

Nucleotide sequence of the BsuRI restriction-modification system.

The genes of the 5'-GGCC specific BsuRI restriction-modification system of Bacillus subtilis have been cloned and expressed in E. coli and their nucleotide sequence has been determined. The restriction and modification genes code for polypeptides with calculated molecular weights of 66,314 and 49,642, respectively. Both enzymes are coded by the same DNA strand. The restriction gene is upstream of the methylase gene and the coding regions are separated by 780 bp. Analysis of the RNA transcripts by S1-nuclease mapping indicates that the restriction and modification genes are transcribed from different promoters. Comparison of the amino acid sequences revealed no homology between the BsuRI restriction and modification enzymes. There are, however, regions of homology between the BsuRI methylase and two other GGCC specific modification enzymes, the BspRI and SPR methylases.

Bacillus subtilis↗