Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.30.2 (endonuclease)
 18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Substrate recognition by the EcoRI restriction endonuclease was investigated by analysis of the nucleotide sequences at the sites of enzymatic cleavage in various DNA molecules. 5'-end labeling and homochromatographic fingerprinting led to the determination of a 17-base-pair sequence spanning the EcoRI site of simian virus 40 DNA and a 15-base-pair sequence overlapping the EcoRI site of Col El plasmid DNA. Three other DNAs were similarly tested, although extended sequences were not determined in these cases. The EcoRI site was shown to be symmetric double-stranded equivalent of -N-G-A-A-T-T-C-N-.
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PMID:Sequences spanning the EcoRI substrate site. 17 27

Limited T1 RNase digestion of subfragments of the SV40 DNA restriction endonuclease fragment EcoRII-G were prepared and analyzed. The fragments were separately labeled with 32P at their 5' terminus and the terminal sequences analyzed with limited snake venom diesterase digestion. The data permitted us to deduce the nucleotide sequence for EcoRII-G. The sequence contains a stretch of 17 A-T base pairs preceding the DNA complementary to the 5' end of "early" message RNA, a stretch of 27 bases with a perfect 2-fold rotational symmetry near the origin of DNA replication and a perfect tandem repeat of 21 nucleotides.
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PMID:Nucleotide sequence of a fragment of SV40 DNA that contains the origin of DNA replication and specifies the 5' ends of "early" and "late" viral RNA. III. Construction of the total sequence of EcoRII-G fragment of SV40 DNA. 18 10

The substrate specificity of the EcoRI restriction endonuclease can be varied in vitro by changing the pH and the ionic environment of the reaction. Phosphodiester bond cleavage occurs at a DNA hexanucleotide sequence d(N-G-A-A-T-T-C-N)/d(N-C-T-T-A-A-G-N) when the ionic strength is high, 100 mM Tris-HCl, 50 mM NaCl, 5 mM MgCl2, and the pH is approximately 7.3. Lowering the ionic strength to 25 mM Tris-HCl, 2 mM MgCl2, and adjusting the pH to 8.5 reduces the recognition specificity of the EcoRI endonuclease to the tetranucleotide sequence, d(N-A-A-T-T-N)/d(N-T-T-A-A-N). The enzymatic activity responsible for this substrate recognition is referred to as EcoRI. Cleavage of pVH51 plasmid DNA under EcoRI conditions results in a number of partial digest fragments, some of which disappear slowly over a prolonged digestion period. This suggests that different recognition sites are cleaved at different rates. Comparison of DNA fragment patterns of modified and unmodified pVH51 DNA indicates that the canonical EcoRI sequence is the most rapidly cleaved site under EcoRI conditions. DNA modified in vivo by the EcoRI methylase is not cleaved by the EcoRI endonuclease under standard conditions, but is cleaved under EcoRI conditions at sites other than the standard EcoRI substrate.
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PMID:Specificity of substrate recognition by the EcoRI restriction endonuclease. 24 1

Superhelical covalently closed circular replicative form DNA (RF I) of coliphage M13 appears as a relaxed molecule that has a base-unpaired region in the form of a bubble (100 to 200 base pairs long) seen in electron micrographs when spread in the presence of formaldehyde and formamide or after pretreatment with glyoxal. S1 endonuclease, specific for single-stranded DNA, converts superhelical M13 RF I DNA, but not nonsuperhelical M13 RF I to a significant extent, into unit-length linear molecules by sequential nicking of two strands. The locations of S1 nuclease-susceptible sites and glyoxal-fixed base-unpaired regions were both related to the five A-T-rich regions in M13 RF DNA. While S1 nuclease does not show preference for any of these sites, glyoxal-fixed bubbles occur predominantly at the major A-T-rich region in M13 RF DNA.
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PMID:Base-unpaired regions in supercoiled replicative form DNA of coliphage M13. 32 5

The recognition sequence for the dam methylase of Escherichia coli K12 has been determined directly by use of in vivo methylated ColE1 DNA or DNA methylated in vitro with purified enzyme. The methylase recognizes the symmetric tetranucleotide d(pG-A-T-C) and introduces two methyl groups per site in duplex DNA with the product of methylation being 6-methylaminopurine. This work has also demonstrated that Dpn I restriction endonuclease cleaves on the 3' side of the modified adenine within the methylated sequence to yield DNA fragments possessing fully base-paired termini. All sequences in ColE1 DNA methylated by the dam enzyme are subject to double strand cleavage by Dpn I endonuclease. Therefore, this restriction enzyme can be employed for mapping the location of sequences possessing the dam modification.
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PMID:Recognition sequence of the dam methylase of Escherichia coli K12 and mode of cleavage of Dpn I endonuclease. 36 70

Chemical syntheses of the two dodecanucleotides d(T-C-A-A-C-G-T-A-A-C-A-C) and d(A-C-G-T-T-G-A-G-A-A-A-G), the two undecanucleotides d(T-T-T-A-C-A-G-C-G-G-C) and d(T-G-T-A-A-A-G-T-G-T-T), the decanucleotide d(A-G-T-C-C-G-A-A-A-G), and the nonanucleotide d(A-A-T-T-C-T-T-T-C) are described. These deoxyribo-oligonucleotide segments, excluding the decanucleotide, represent the DNA duplex corresponding to the previously determined nucleotide sequence -30 to -51 of the promoter region of the gene for the tyrosine suppressor tRNA (Sekiya, T., Gait, M.J., Norris, K., Ramamoorthy, B., and Khorana, H.G. (1976) J. Biol. Chem. 251, 4481-4489) and include the EcoRI restriction endonuclease sequence at the appropriate 5'-end. The nona- and decanucleotide along with the previously synthesized deoxyribo-oligonucleotide segments 25 to 27 (Ramamoorthy, B., Lees, R.G., Kleid, D., and Khorana, H.G. (1976) J. Biol. Chem. 251, 676-694) together represent the DNA duplex corresponding to the natural nucleotide sequence 121 to 142 of the region adjoining the C-C-A end of the tyrosine tRNA gene and, in addition, a run of nine nucleotides which include the EcoRI restriction enzyme sequence at the 5'-end. The syntheses used protected mono- and oligonucleotides and stepwise condensation methods. A noteworthy feature of the present syntheses was the use of reverse phase high pressure liquid chromatography for the rapid and efficient separation of synthetic reaction mixtures.
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PMID:Total synthesis of a tyrosine suppressor transfer RNA gene. XIV. Chemical synthesis of oligonucleotide segments corresponding to the terminal regions. 37 18

The nucleotide sequence A-A-T-T was inserted into the intergenic region of the f1 genome at a site cleaved by Hae III (cleavage sequence (G-G-C-C). The resultant viable phage mutant (R199) contains a single site sensitive to the restriction endonuclease EcoRI (cleavage sequence G-A-A-T-T-C). This phage is sensitive to EcoRI restriction and modification in vivo and in vitro. Its potential for use as a cloning vector has been tested by construction in vitro of an f1/pBR322 chimeric phage. The four bases inserted into wild-type f1 to generate the R199 mutant came from a small restriction fragment obtained by digesting plasmid pBR322 with EcoRI and HindIII. The use of this linker prepared from a biological substrate is an example of a technique for constructing restriction enzyme sites in vitro. It is presented as an alternative to the use of synthetic linkers and should be generally applicable.
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PMID:Insertion mutant of bacteriophage f1 sensitive to EcoRI. 37 63

In studies of some sequence dependent structural factors and stabilizing effects of oligonucleotides the octanucleotide d(pG-A-T-C-T-T-T-T) was of particular interest in view of the presence of an endonuclease cleavage site. Its chemical synthesis is reported as well as the structural effects in CD spectral properties of the octanucleotide and of some related compounds.
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PMID:Conformational properties of the octanucleotide d(pG-A-T-C-T-T-T-T) and its intermediates. 42 13

The base sequence of mouse L-cell satellite DNA was investigated by degradation of the two separated complementary strands with the base specific enzyme, T4 endonuclease IV. Digestion of the heavy strand DNA released a limited number of oligonucleotides which were separated by ionophoresis/homochromatography, isolated, and sequenced by the 'wandering spot' method. The light strand DNA was resistant to digestion with T4 endonuclease IV and no detectable amounts of oligonucleotides were released. The oligonucleotides obtained from the heavy strand were related in sequence, indicating that mouse satellite DNA derived from a short tandemly repeated sequence. The sequence of part of the original repeat unit is proposed to be C-A-T-T-T-T-T-C. Five major oligonucleotides were identified, all of which differ from the proposed original sequence by single base changes. The five major oligonucleotides occur with about equal frequency and together comprise approximately 50% of the oligonucleotides released by T4 endonuclease IV from the heavy strand DNA. In addition to the five major oligonucleotides, several oligonucleotides were found to occur in lesser amounts. Since these oligonucleotides are related to the major oligonucleotides, it is likely that they have arisen from them by mutation.
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PMID:Sequence studies on mouse L-cell satellite DNA by base-specific degradation with T4 endonuclease IV. 71 12

The nucleotide sequence of a double stranded DNA fragment from the gene AB region of bacteriophage S13 DNA has been determined. The fragment was isolated as two adjacent shorter fragments by cleavage of S13 replicative form (RF) DNA with restriction endonuclease III from Hemophilus aegyptius. The strands of the fragments were separated electrophoretically and hydrolyzed with T4 endonuclease IV to yield short oligonucleotides which were then sequenced by partial exonuclease digestion. The complete nucleotide sequence of the restriction fragments was obtained by ordering the inter- and intrastrand overlapping oligonucleotide sequences. The adjacent fragments were 190 nucleotides in length. The sequences included a HindII site, an AluI site and two sequences which may be possible transcription initiation sequences, one with an adjacent sequence homologous to the canonical promoter site sequence T-A-T-Pu-A-T-Pu. Examination of the three possible reading frames for translation of the sequence revealed only one possible complete translation product. The postulated partial sequence of gene A protein has a highly positively charged arginine-rich area which may have importance in DNA binding.
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PMID:The nucleotide sequence of two restriction fragments located in the gene AB region of bacteriophage S13. 90 72


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