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)

Overlapping clones of the structural gene region for alpha-amylase, Amy, were isolated from a lambda EMBL4 library containing genomic DNA fragments from an amylase-null strain of Drosophila melanogaster. Southern blot analysis and restriction endonuclease mapping of the cloned region indicate that it contains an Amy gene duplication within an inverted repeat sequence as is characteristic of the genomic arrangement for this species. Spacing between the cloned gene copies is similar to that commonly found in other strains. Evidence is presented for the presence of an inversion 4 to 9 kb in length within the cloned Amy region of the null strain. We postulate a causal relationship between the presence of the inversion and the failure of individuals from the null strain to express amylase. A model is proposed that suggests the inversion may have arisen through intramolecular (or sister-strand) recombination mediated by homologous pairing of the inverted repeat sequences at the Amy locus.
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PMID:Molecular cloning of alpha-amylase genes from Drosophila melanogaster. III. An inversion at the Amy locus in an amylase-null strain. 249 82

We have identified an endonuclease(s) that preferentially cleaves the internucleosomal linker regions in the aleurone chromatin producing mono- and oligonucleosomes. This enzyme(s) has been designated as a "linker"-specific nuclease(s). This nuclease does not require divalent cations for activity, and therefore it is not the "Ca2+-Mg2+-DNase" found in mammalian cells. The linker-specific nuclease activity is not detectable in the dry aleurone tissue and in the tissue treated with 0.5 mM cordycepin. The endonuclease activity of the aleurone tissue incubated with gibberellic acid is higher than the level of this endonuclease in tissue treated with abscisic acid or water alone. Nuclei isolated from embryos have lower levels of endonuclease activities compared to those from aleurone tissue. Digestion of the nuclei from embryos with micrococcal nuclease revealed the subunit structure of chromatin. In Southern blots of the HindIII digests of DNA from embryos, five DNA bands hybridized to a nick-translated alpha-amylase cDNA clone. In similar autoradiograms with aleurone DNA, particular bands are less visible, notably in the DNA isolated from the tissue treated with gibberellic acid. This is the first report of the presence of a linker-specific nuclease activity in plant cells.
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PMID:Study of barley endonucleases and alpha-amylase genes. 301 70

The gene coding for alpha-amylase from Bacillus amyloliquefaciens was isolated by direct shotgun cloning using B. subtilis as a host. The genome of B. amyloliquefaciens was partially digested with the restriction endonuclease MboI and 2- to 5-kb fragments were isolated and joined to plasmid pUB110. Competent B. subtilis amylase-negative cells were transformed with the hybrid plasmids and kanamycin-resistant transformants were screened for the production of alpha-amylase. One of the transformants producing high amounts of alpha-amylase was characterized further. The alpha-amylase gene was shown to be present in a 2.3-kb insert. The alpha-amylase production of the transformed B. subtilis could be prevented by inserting lambda DNA fragments into unique sites of EcoRI, HindIII and KpnI in the insert. Foreign DNA inserted into a unique ClaI site failed to affect the alpha-amylase production. The amount of alpha-amylase activity produced by this transformed B. subtilis was about 2500-fold higher than that for the wild-type B. subtilis Marburg strain, and about 5 times higher than the activity produced by the donor B. amyloliquefaciens strain. Virtually all of the alpha-amylase was secreted into the culture medium. The secreted alpha-amylase was shown to be indistinguishable from that of B. amyloliquefaciens as based on immunological and biochemical criteria.
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PMID:Molecular cloning of alpha-amylase gene from Bacillus amyloliquefaciens and its expression in B. subtilis. 618 69

The gene for thermostable alpha-amylase from the thermophilic bacterium Bacillus stearothermophilus has been cloned and expressed in Escherichia coli. Each alpha-amylase-producing colony contained at least a 9.7-kilobase-pair (kb) chimeric plasmid composed of the vector pBR322 and a common 5.4-kb HindIII fragment of DNA. B. stearothermophilus contains four plasmids with sizes from 12 kb to over 108 kb. Restriction endonuclease analysis of these naturally occurring plasmids showed they also contain a 5.4-kb HindIII fragment of DNA. Cloning experiments with the four plasmids yielded alpha-amylase-producing E. coli that contained the same 9.7-kb chimeric plasmid. Restriction endonuclease analysis and further recombinant DNA experiments identified a 26-kb plasmid that contains the gene for alpha-amylase. A spontaneous mutant of B. stearothermophilus unable to produce alpha-amylase was missing the 26-kb plasmid but contained a 20-kb plasmid. A 6-kb deletion within the region of the 5.4-kb HindIII fragment yielded the 20-kb plasmid unable to code for alpha-amylase. A nick-translated probe for the alpha-amylase coding region did not hybridize to either plasmid or total cellular DNA from this mutant strain of B. stearothermophilus. These results demonstrate the gene for alpha-amylase is located exclusively on a 26-kb plasmid in B. stearothermophilus with no genetic counterpart present on the chromosome.
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PMID:Bacillus stearothermophilus contains a plasmid-borne gene for alpha-amylase. 619 26

We have chemically synthesized an oligonucleotide 5'd(TGATTGATTGA)3' 3'd(ACTAACTAACT)5' that encodes the translation termination codon TGA in all three reading frames. After ligation of appropriate restriction endonuclease linkers to the ends, the double-stranded oligonucleotide (STOP-oligonucleotide) was joined to the plasmid pBR322 between the EcoRI and BamHI, or HindIII and BamHI sites, and the hybrid plasmids were transformed into Escherichia coli HB101. Four different constructions were obtained: (i) EcoRI-STOP-BamHI (STOP-oligonucleotide flanked by EcoRI and BamHI linkers; pKTH606), (ii) HindIII-STOP-BamHI (pKTH601), (iii) BamHI-STOP-HindIII (pKTH604), and (iv) HindIII-STOP-POTS-BamHI (two STOP-oligonucleotides in opposite orientation; pKTH605). The inserts in pKTH606 and pKTH601 were excised and transferred to a modified plasmid constructed previously for the expression and secretion of foreign gene products from Bacillus subtilis. The resulting secretion plasmids now contain the promoter/signal sequence region of the alpha-amylase gene from Bacillus amyloliquefaciens joined to the STOP-oligonucleotide by EcoRI or HindIII linkers. Foreign genes can be cloned into these sites. The plasmids can be used to express foreign genes truncated at their C-terminal end and therefore lacking their own translation termination codon. One such plasmid has been successfully used to express the Semliki Forest virus (SFV) membrane protein E1 truncated at its C-terminus.
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PMID:Chemical synthesis and molecular cloning of a STOP oligonucleotide encoding an UGA translation terminator in all three reading frames. 631 80

Functional analysis of a gibberellin-regulated wheat alpha-amylase promoter, alpha-Amy2/54, has indicated that three regions were essential for expression. By studying the ability of mutant promoters, containing a randomly inserted 22 bp excision linker, to direct expression in oat aleurone protoplasts we have refined the positions and extents of these three cis elements and also demonstrated the presence of two additional elements. By converting the linker insertions to either single base point mutations or deletions using the class IIS restriction endonuclease BsmI we have shown that nucleotides -119 and -109 within the GARE -121GTAACAGAGTCTGG-108 and nucleotide -152 within the proposed element -156GATTGACTTGACC-144 are essential for high level expression from this promoter.
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PMID:Functional analysis of linker insertions and point mutations in the alpha-Amy2/54 GA-regulated promoter. 854 1

When Bacillus subtilis levanase (SacC), alpha-amylase (AmyE) and chitosanase (Csn) structural genes were expressed under the regulated control of sacR, the inducible levansucrase (SacB) leader region in a degU32(Hy) mutant, it was observed that the production yields of the various extracellular proteins were quite different. This is mainly due to differences in the stabilities of their corresponding mRNAs which lead to discrepancies between the steady-state level of mRNA of sacB and csn on the one hand and amyE and sacC on the other. In contrast to levansucrase mRNA, the decay curves of alpha-amylase and levanase mRNAs obtained by Northern blotting analysis did not match the decay curves of their functional mRNA. This suggested that only a part of the population of the amyE and sacC transcripts was fully translated, while the others were possibly poorly bound to ribosomes and thus were only partially translated or not at all and consequently submitted to rapid endonuclease degradation. This hypothesis was substantiated by the finding that the introduction of a Shine-Dalgarno sequence upstream from the ribosome-binding site in the sacC transcript resulted in a fourfold increase in both the half-life of this transcript and the production of levanase. An additional cause of low-level levanase production is the premature release of mRNA by the polymerase. It was attempted to correlate this event with internal secondary structures of sacC mRNA.
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PMID:Transcripts of the genes sacB, amyE, sacC and csn expressed in Bacillus subtilis under the control of the 5' untranslated sacR region display different stabilities that can be modulated. 1132 Jan 36

The purpose of this study was to examine the genetic structure of the typical commensal Streptococcus mitis biovar 1 in its natural habitat in the human oral cavity and pharynx and to investigate the role that selected microbial properties and host, spatial, and temporal factors play in determining the structure of the bacterial population. Consecutive samples were collected from buccal and pharyngeal mucosal surfaces of two infants, their four parents, and two elderly individuals over a period of approximately 1 year. A total of 751 isolates identified as S. mitis biovar 1 were typed by restriction endonuclease analysis (REA) and representative clones were typed by multilocus enzyme electrophoresis (MLEE). The genetic diversity of the S. mitis biovar 1 isolates collected from single infant hosts over a period of 9 to 10 months was found to be between 0.69 and 0.76, which is considerably higher than that previously observed for intestinal populations of Escherichia coli. The study provides evidence of the existence of both transient and persistent clones in adult individuals. In the two infants, however, none of 42 demonstrated clones were detected on more than a single occasion. Statistical calculations showed that the ability to persist was not distributed at random in the S. mitis biovar 1 population. However, neither immunoglobulin A1 protease activity nor the ability to bind alpha-amylase from saliva was a preferential characteristic of persistent genotypes. In contrast to current concepts of climax ecosystems, the species niche in the habitat appears to be maintained predominantly by a succession of clones rather than by stable strains. Several lines of evidence suggest that the major origin of "new" clones is the many other habitats in the respiratory tract that are occupied by this species.
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PMID:Population dynamics of Streptococcus mitis in its natural habitat. 1155 43