Characterization of the cloned BamHI restriction modification system: its nucleotide sequence, properties of the methylase, and expression in heterologous hosts

Brooks, Joan E.; Nathan, Peter D.; Landry, David; Sznyter, Laura A.; Waite-Rees, Phyllis A.; Ives, Catherine L.; Moran, Laurie S.; Slatko, Barton E.; Benner, Jack S.

doi:10.1093/nar/19.4.841

Cited by 63 publications

(40 citation statements)

References 64 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Plasmid pACYC184 was used as a control in several complementation experiments (9). Plasmid pBamRM2.7-HP13, containing the complete BamHI system, was described previously (8). pY4Cm, a derivative of pGEM-Blue (Promega Biotec), was used to construct clones that would integrate into the B. subtilis chromosome (17).…”

Section: Methodsmentioning

confidence: 99%

“…BamHI was one of the first RM systems discovered (37), and its endonuclease (R.BamHI) and methylase (M.BamHI) have been purified to apparent homogeneity and characterized (8,18,23,30). The BamHI RM system has been cloned and the nucleotide sequence determined; a small open reading frame encoding 102 amino acids located between the methylase and endonuclease genes was discovered (8). This open reading frame was previously shown to regulate bamHIM expression (8,24) and has a predicted amino acid sequence very similar to that of pvuIIC (34).…”

mentioning

confidence: 99%

“…The BamHI RM system has been cloned and the nucleotide sequence determined; a small open reading frame encoding 102 amino acids located between the methylase and endonuclease genes was discovered (8). This open reading frame was previously shown to regulate bamHIM expression (8,24) and has a predicted amino acid sequence very similar to that of pvuIIC (34). It has been designated bamHIC (for BamHI controlling element).…”

mentioning

confidence: 99%

See 2 more Smart Citations

Regulation of the BamHI restriction-modification system by a small intergenic open reading frame, bamHIC, in both Escherichia coli and Bacillus subtilis

1992

View full text Add to dashboard Cite

BamHI, from Bacillus amyloliquefaciens H, is a type II restriction-modification system recognizing and cleaving the sequence G--GATCC. The BamHI restriction-modification system contains divergently transcribed endonuclease and methylase genes along with a small open reading frame oriented in the direction of the endonuclease gene. The small open reading frame has been designated bamHIC (for BamHI controlling element). It acts as both a positive activator of endonuclease expression and a negative repressor of methylase expression of BamHI clones in Escherichia coli. Methylase activity increased 15-fold and endonuclease activity decreased 100-fold when bamHIC was inactivated. The normal levels of activity for both methylase and endonuclease were restored by supplying bamHIC in trans. The BamHI restriction-modification system was transferred into Bacillus subtilis, where bamHIC also regulated endonuclease expression when present on multicopy plasmid vectors or integrated into the chromosome. In B. subtilis, disruption of bamHIC caused at least a 1,000-fold decrease in endonuclease activity; activity was partially restored by supplying bamHIC in trans.

show abstract

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

See 1 more Smart Citation

Regulation of the BamHI restriction-modification system by a small intergenic open reading frame, bamHIC, in both Escherichia coli and Bacillus subtilis

1992

View full text Add to dashboard Cite

show abstract

“…The coding sequence for BamHI has been cloned and sequenced, and the enzyme has been overexpressed in Escherichia coli (2,3,10). Like all type II restriction endonucleases, BamHI requires the divalent cation Mg2' as a cofactor for cleavage activity.…”

mentioning

confidence: 99%

Cofactor requirements of BamHI mutant endonuclease E77K and its suppressor mutants

Schildkraut

1991

J Bacteriol

View full text Add to dashboard Cite

A mutant BamHI endonuclease, E77K, belongs to a class of catalytic mutants that bind DNA efficiently but cleave DNA at a rate more than 103-fold lower than that of the wild-type enzyme (S. Y. Xu and I. Schildkraut, J. Biol. Chem. 266:4425-4429, 1991). The preferred cofactor for the wild-type BamHI is Mg2+. BamHI is 10-fold less active with Mn2+ as the cofactor. In contrast, the E77K variant displays an increased activity when Mn2+ is substituted for Mg2, in the reaction buffer. Mutations that partially suppress the E77K mutation were isolated by using an Escherirhia coli indicator strain containing the dinD::lacZ fusion. These pseudorevertant endonucleases induce E. coli SOS response (as evidenced by blue colony formation) and thus presumably nick or cleave chromosomal DNA in vivo. Consistent with the in vivo result, the pseudorevertant endonucleases in the crude cell extract display site-specific partial DNA cleavage activity. DNA sequencing revealed two unique suppressing mutations that were 'located within two amino acid residues of the original mutation. Both pseudorevertant proteins were purified and shown to increase specific activity at least 50-fold. Like the wild-type enzyme, both pseudorevertant endonucleases prefer Mg2Y as the cofactor. Thus, the second-site mutation not only restores partial cleavage activity but also suppresses the metai preference a$ well. These results suggest that the Glu-77 residue may play a role in metal ion binding or in enzyme activation (allosteric transition) following sequence-specific recognition.The BamHI restriction endonuclease purified from Bacillus amyloliquefaciens H cleaves the symmetric DNA sequence 5'-GGATCC-3' between the two guanines on both strands (reviewed in reference 17). The coding sequence for BamHI has been cloned and sequenced, and the enzyme has been overexpressed in Escherichia coli (2,3,10). Like all type II restriction endonucleases, BamHI requires the divalent cation Mg2' as a cofactor for cleavage activity. Mn2+ can be substituted for Mg2+ in the reaction, but the enzyme activity is reduced (24). Mn2+ also decreases the enzyme specificity, allowing BamHI to cleave noncanonical sequences (BamHI star sites; 10, 14). The DNA-binding activity, however, is independent of metal cofactor (17, 28).The SOS regulon consisting of about 20 genes in E. coli is negatively regulated by the LexA repressor (reviewed in references 19, 22, and 25). The repressor can be cleaved and inactivated by the RecA protein to derepress the genes in the regulon. The proteolytic activity of the RecA protein is activated by a signal (e.g., single-stranded DNA or oligonucleotides) upon DNA damage or inhibition of DNA replication. As the result of DNA damage or DNA replication interference, the genes in the SOS regulon are expressed at an increased level. E. coli indicator strain AP1-200 contains a dinD: :lacZ fusion in which the lactose operon is fused to a DNA damage-inducible promoter (12). --Galactosidase expression is increased upon treatment of the cells with UV, mitomycin...

show abstract

“…The gene encoding the BamHI endonuclease has previously been cloned (11), sequenced (12), and its product has been overexpressed and crystallized (13). We are pursuing the genetic determination of the residues in BamHI that are involved in catalysis and recognition, while the three dimensional structure is being determined (14).…”

Section: Introductionmentioning

confidence: 99%

Direct selection of binding proficient/catalytic deficient variants ofBamHI endonuclease

Dorner

Schildkraut

1994

Nucl Acids Res

View full text Add to dashboard Cite

Variants of BamHl endonuclease in which the glutamate 1 13 residue has been changed to lysine or the aspartate 94 to asparagine were shown to behave as repressor molecules in vivo. This was demonstrated by placing a BamHl recognition sequence, GGATCC, positioned as an operator sequence in an antisense promoter for the aadA gene (spectinomycin resistance). Repression of this promoter relieved the inhibition of expression of spectinomycin resistance. This system was then used to select new binding proficient/cleavage deficient BamHl variants. The BamHl endonuclease gene was mutagenized either by exposure to hydroxylamine or by PCR. The mutagenized DNA was reintroduced into E.coli carrying the aadA gene construct, and transformants that conferred spectinomycin resistance were selected. Twenty Spr transformants were sequenced. Thirteen of these were newly isolated variants of the previously identified D94 and E113 residues which are known to be involved in catalysis. The remaining seven variants were all located at residue 111 and the glutamate 111 residue was shown to be involved with catalysis.

show abstract

Characterization of the cloned BamHI restriction modification system: its nucleotide sequence, properties of the methylase, and expression in heterologous hosts

Cited by 63 publications

References 64 publications

Regulation of the BamHI restriction-modification system by a small intergenic open reading frame, bamHIC, in both Escherichia coli and Bacillus subtilis

Regulation of the BamHI restriction-modification system by a small intergenic open reading frame, bamHIC, in both Escherichia coli and Bacillus subtilis

Cofactor requirements of BamHI mutant endonuclease E77K and its suppressor mutants

Direct selection of binding proficient/catalytic deficient variants ofBamHI endonuclease

Contact Info

Product

Resources

About