PubMed Health⌕ Search

Biomedical subjects

V M Kramarov

Publications and source records attributed to V M Kramarov.

At least 19 recordsLinked to original sources
PubMed1999

Mutation S543N in the thumb subdomain of the Taq DNA polymerase large fragment suppresses pausing associated with the template structure.

Substitution of Asn for the conserved Ser543 in the thumb subdomain of the Taq DNA polymerase large fragment (Klentaq DNA polymerase) prevents pausing during DNA synthesis and allows the enzyme to circumvent template regions with a complex structure. The mutant enzyme (KlentaqN DNA polymerase) provides specific PCR amplification and sequencing of difficult templates, e.g. those with a high GC% content or strong secondary structure.

Asparagine↗
PubMed1997

[DNA polymerase mediated amplification of DNA fragments using primers with mismatches in the 3'-region].

The ability of three thermostable enzymes, Tth, Taq, and Klentaq DNA polymerases, to amplify DNA with primers containing mismatches in the 3'-terminal region was studied. It is shown that Tth polymerase, in contrast to the Taq and Klentaq enzymes, synthesizes equally well DNA with primers perfectly complementary to the template and with those containing mismatches next the 3'-end. The use of Tth DNA polymerase in the polymerase chain reaction was shown to result, in some cases, in a great number of additional, nonspecific DNA fragments as compared with Taq DNA polymerase. This may be due to the ability of Tth polymerase for DNA primer extension even if the 3'-terminal region of the primer contains nucleotides non-complementary to the template. Tth DNA polymerase and a Klentaq/Tth mixture (100:1) can be efficiently used in the amplification of DNA with degenerated primers and primers forming nonperfect duplexes with the template.

DNA↗
PubMed1993

[Isolation of site-specific endonucleases and methylases from Bacillus cereus 83].

The site-specific endonuclease R Bce83I and methylase M Bce83I were isolated from Bacillus cereus 83 by three consecutive chromatographies on blue agarose, hydroxyapatite and heparin-Sepharose. R Bce83I recognizes [formula: see text] sequences on the DNA and cleaves the DNA as indicated by the arrows. The endonuclease is stimulated by S-adenosyl-L-methionine and may consequently be referred to type IV restriction enzymes.

Bacillus cereus↗
PubMed1993

[New site site-specific endonuclease and methylase from Bacillus licheniformis 736].

The site-specific endonuclease R Bli736I and methylase M Bli736I have been isolated from the Bacillus licheniformis strain 736 by blue-agarose, hydroxyapatite-Ultragel and heparin-Sepharose chromatography. The enzymes are free from interfering impurities. R Bli736I recognizes the 5'-GGTCTCN-3' decreases and decreases 5'-NNNNNGAGACC-3' sequences on the DNA and cleaves the DNA as indicated by arrows to form single-stranded 4-nucleotide 5'-protruding termini. This enzyme is an isoschizomer of Eco3II isolated from E. coli.

Bacillus↗
PubMed1990

Amplification of DNA sequences of Epstein-Barr and human immunodeficiency viruses using DNA-polymerase from Thermus thermophilus.

Using thermophilic DNA-polymerase from Thermus thermophilus we have amplified by polymerase chain reaction (PCR) specific DNA sequences of Epstein-Barr virus (EBV) and human immunodeficiency virus (HIV). DNA-polymerase from Thermus thermophilus (molecular mass of 80-86 kDa) differs in its physico-chemical properties from DNA-polymerase from Thermus aquaticus (molecular mass of 62-68 kDa). To amplify the specific EBV DNA sequence, oligonucleotide primers for the virus replicon region (oriP region) were used. As a result of amplification, a specific 405-bp DNA fragment was produced.

Base Sequence↗
PubMed1989

[New producers of site-specific endonucleases from microorganisms of the Bacillus genus].

52 strains of Bacillus generum have been tested for production of site-specific endonucleases. The sequence recognized by the enzyme was determined for 23 enzymes, the cleavage site inside the sequence was determined for 5 enzymes. All the enzymes under study were found to be isomers of the known enzymes. The selected strains are peculiar for the high level of site-specific endonucleases content and may be used as producents of the enzymes.

Bacillus↗
PubMed1988

[Restriction endonucleases from Bifidobacteria].

The sitespecific restriction endonucleases were found in four strains among the twelve strains of anaerobic bacteria of generum Bifidobacterium. Two of the restriction endonucleases studied, BadI from B. adolescentis LVA1 and BbfI from B. bifidum LVA3, are isoshizomers of XhoI and recognize the nucleotide sequence CTCGAG. The restriction endonucleases Bbf7411I from B. bifidum 7411 and Bla7920I from B. lactentis 7920 recognize and hydrolize the nucleotide sequence TCCGGA having the specifity analogous to the one of restriction endonuclease CauB3I. Like CauB3I, these restriction endonucleases are unable to hydrolyize DNA if the adenine residues in the recognition site are methylated.

Base Sequence↗
PubMed1988

[A new type of cleavage of the recognition site by the site-specific endonuclease Bst 4.4I from Bacillus stearothermophilus 4.4].

A site-specific endonuclease Bst 4.4I was isolated from the cell extract of Bacillus stearothermophilus 4.4 and partially purified by chromatography on Ultragel AcA-44 and heparin-Sepharose. It was shown that the endonuclease cleaves lambda and M13 DNA yielding distinct fragments just as endonucleases of II and III types but, in contrast to them can produce two two-strand cuts separated with 30 to 32 nucleotides in the region of the recognition site.

Base Sequence↗
PubMed1987

Isolation and some properties of the site-specific endonuclease and methylase Bme2161 from Bacillus megaterium 216.

The site-specific endonuclease Bme2161 was isolated as a homogeneous preparation by chromatography on phosphocellulose, hydroxyapatite and heparin-agarose. The molecular mass of the enzyme, determined by gel filtration and by electrophoresis under denaturing conditions, was found to be 60 kDa and 30 kDa respectively. These data indicate that the native enzyme consists of two identical subunits. The enzyme recognized the decreases pentanucleotide sequence 5'-GGACC-3' X 3'-CCTGG-5' and cleaves the sequence as indicated by arrows. The increases optimal concentration for endonuclease reaction is 6-7 mM Mg2+. The endonuclease relaxes its specificity in the presence of glycerol or dimethyl sulfoxide at low Mg2+ concentration (1-3 mM). Methylase Bme2161, which protects DNA against endonuclease Bme2161 action by DNA methylation, was isolated from the same bacterial strain.

Bacillus megaterium↗
PubMed1987

[Site-specific endonuclease CauB31 from Chloroflexus aurantiacus B3].

A sequence-specific endonuclease CauB3I has been isolated from cell extracts of Chloroflexus aurantiacus and partially purified by chromatography on heparin-sepharose; the yield was 3000 units per 1 g of cells. The final preparation is free of non-specific nucleases. It is shown that endonuclease CauB3I recognizes 5' T decreases CCGGA 3' sequence in double-stranded DNA and cleaves it as shown by an arrow. Methylation of adenine in the recognition sequence makes it resistant to CauB3I.

Base Sequence↗
PubMed1983

[Site-specific endonuclease BmeI from Bacillus megaterium 216].

A site-specific endonuclease BmeI has been isolated from Bacillus megaterium 216 by gel filtration on ultragel AcA-44 with a subsequent chromatography on heparin-sepharose 6B. On the double-stranded DNA the endonuclease recognizes the pentanucleotide sequence (Formula: see text); and hydrolyzes it in the points shown by arrows. At gel filtration the endonuclease is eluted in the volume corresponding to a molecular mass of 60 000.

Bacillus megaterium↗
PubMed1983

[BbvII--a new site-specific endonuclease from Bacillus brevis 80].

BbvII, a new site-specific endonuclease, has been isolated from Bacillus brevis 80 by gel-filtration and chromatography on heparin-Sepharose. The endonuclease recognizes a non-symmetrical sequence 5'-GTCTTC-3' in double-stranded DNA and cleaves DNA 3'-CAGAAG-5' in both strands outside the recognition sequence.

Bacillus↗
PubMed1981

[DNA-methylase from Arthrobacter luteus screens DNA from the action of site-specific endonuclease Alu I].

DNA-methylase was isolated from a cell extract of A. luteus and partially purified by chromatography on phosphocellulose. The purified enzyme methylates DNA of phage lambda and plasmids pBr322, thus making them resistant to a subsequent action of endonuclease Alu I. It has been shown that cytosine is the object of methylation within DNA. This modification does not screen DNA from the action of site-specific endonucleases Sal I, Bam HI, EcoR I, EcoR II, Xho I and Xho II. It has been experimentally demonstrated that the isolated methylase is site-specific and identifies in the DNA the nucleotide sequence 5'-AGCT-3', by methylating cytosine in the DNA.

Arthrobacter↗