Streptomyces aureofaciens B96 produces several intra- and extracellular enzymes with deoxyribonuclease activity. According to the sequence of the previously published gene exoSc from S. coelicolor A3(2), the DNA sequence from S. aureofaciens B96 was amplified, cloned and expressed in E. coli. The protein product of exoSa gene, recExoSa, was also an exonuclease with DNAase and 5'-phosphomonoesterase activities at optimum temperature 37 degrees C and pH 8.0. It degraded only linear DNA (chromosomal, double-stranded and single-stranded) and linear plasmid DNA from both ends, with a preference to blunt ends in comparison with overhang ends. The purified enzyme exhibited no RNAase activity. Both exoSc and exoSa genes were interrupted by the apramycin resistance gene; constructed fragments were transformed into particular streptomyces protoplasts. Mutation caused by exoSa disruption in S. aureofaciens chromosome and mutation by interrupted exoSc in S. coelicolor were lethal.

• MeSH
• bakteriální proteiny chemie   genetika   izolace a purifikace   metabolismus   MeSH
• deoxyribonukleasy chemie   genetika   izolace a purifikace   metabolismus   MeSH
• exonukleasy chemie   genetika   izolace a purifikace   metabolismus   MeSH
• fosfatasy chemie   genetika   izolace a purifikace   metabolismus   MeSH
• klonování DNA MeSH
• molekulární sekvence - údaje MeSH
• sekvence aminokyselin MeSH
• sekvenční seřazení MeSH
• stabilita enzymů MeSH
• Streptomyces aureofaciens chemie   enzymologie   genetika   MeSH
• Streptomyces coelicolor chemie   enzymologie   genetika   MeSH
• substrátová specifita MeSH
• umlčování genů * MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• bakteriální proteiny MeSH
• deoxyribonukleasy MeSH
• exonukleasy MeSH
• fosfatasy MeSH

Tetracycline-producing strains of Streptomyces aureofaciens expressed SauLPI restriction-modification (R-M) system, which recognized specific DNA sequence 5'-GCCGGC-3' (isoschizomer Nael). The activation of the second R-M system SauLPII (5'-GAGCTC-3', isoschizomer of XhoI), which was silent during the growth cycle, after a foreign DNA transfer into this strain was observed. This phenomenon was tentatively explained as a response of the cells against the exogenous DNA entering the cells. The involvement of a SOS-like response in induction of R-M system genes in S. aureofaciens strains has been considered.

• MeSH
• bakteriofág mu genetika   MeSH
• replikace DNA MeSH
• restrikční endonukleasy typu II metabolismus   MeSH
• SOS odpověď (genetika) MeSH
• Streptomyces aureofaciens enzymologie   genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• CTCGAG-specific type II deoxyribonucleases MeSH Prohlížeč
• restrikční endonukleasy typu II MeSH

A Streptomyces aureofaciens gene, gap, encoding glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was previously identified. Hybridization studies suggested the presence of a second gap gene in S. aureofaciens. To clone the gene, S. aureofaciens subgenomic library was screened with an oligonucleotide probe encoding a peptide motif conserved in all GAPDH. 3352 bp positive BamHI fragment was identified, the length of which correlated with the hybridization signal. The nucleotide sequence of the fragment was determined, and analysis of the sequence revealed the presence of three open reading frames (ORF). However, none of the genes coded for GAPDH. All three genes formed an operon, consisting of gene orf251, with a high homology to a conserved gene present only in archaeabacteria, and the aldA and adhA genes homologous to various eukaryotic and prokaryotic aldehyde- and alcohol-dehydrogenases, with maximum homology to the phenylacetaldehyde dehydrogenases and arylalcohol dehydrogenases, respectively.

• MeSH
• aldehyddehydrogenasa chemie   genetika   metabolismus   MeSH
• klonování DNA * MeSH
• molekulární sekvence - údaje MeSH
• sekvence aminokyselin MeSH
• sekvence nukleotidů MeSH
• sekvenční analýza DNA MeSH
• sekvenční seřazení MeSH
• Streptomyces aureofaciens enzymologie   genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• aldehyddehydrogenasa MeSH

The first valine dehydrogenase of S. aureofaciens had been described (Vancurová, I., Vancura, A., Volc, J., Neuzil, J., Flieger, M., Basarová, G. and Bĕhal, V. (1988) J. Bacteriol. 170, 5192-5196). In the present work, a second valine dehydrogenase was detected and purified by hydrophobic and fast protein liquid chromatographies. The enzyme has a relative molecular mass (M(r)) of 240,000 and is composed of 6 identical subunits, each of M(r) 41,000. In the presence of NAD, the enzyme catalyzes the reversible deamination of several branched- and straight-chain amino acids. The enzyme activities with L-2-aminobutyrate and deamino-NAD+ are markedly higher than those with L-valine and NAD+, respectively. The enzyme synthesis is significantly induced by L-valine but severely repressed by ammonia. Molecular and catalytic properties of the enzyme distinguish it from the other described valine dehydrogenases. The results directly demonstrate the presence of two valine dehydrogenases in a single Streptomyces species.

• MeSH
• koncentrace vodíkových iontů MeSH
• oxidoreduktasy aminokyselin antagonisté a inhibitory   chemie   izolace a purifikace   MeSH
• Streptomyces aureofaciens enzymologie   MeSH
• substrátová specifita MeSH
• teplota MeSH
• valindehydrogenasa (NADP+) MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• oxidoreduktasy aminokyselin MeSH
• valindehydrogenasa (NADP+) MeSH
• valine dehydrogenase (NAD+) MeSH Prohlížeč

A novel gene (hur) conferring resistance to hydroxyurea (HU) in Escherichia coli has been identified in a Streptomyces aureofaciens genomic library. The expression of hur in E. coli was under the control of the external plasmid tet promoter. Sequence analysis of a minimal fragment revealed an open reading frame (ORF) encoding a protein of 340 amino acids with an M(r) of 36,049 and an average hydropathy index of 1.13. The predicted protein product was similar to streptomycin phosphotransferases from Streptomyces glaucescens and Streptomyces griseus (52.4% and 50.8% identity, respectively), but it did not confer resistance to streptomycin or to any of the other aminoglycoside antibiotics tested. It is inferred that hur encodes a phosphotransferase that inactivates HU by phosphorylation of the hydroxy group in the hydroxylamine moiety.

• MeSH
• antibiotická rezistence genetika   MeSH
• bakteriální geny MeSH
• Escherichia coli genetika   MeSH
• fosfotransferasy s alkoholovou skupinou jako akceptorem * MeSH
• fosfotransferasy chemie   genetika   MeSH
• hydroxymočovina farmakologie   MeSH
• molekulární sekvence - údaje MeSH
• rekombinantní DNA genetika   MeSH
• restrikční mapování MeSH
• ribonukleotidreduktasy genetika   MeSH
• sekvence aminokyselin MeSH
• sekvence nukleotidů MeSH
• sekvenční homologie nukleových kyselin MeSH
• Streptomyces aureofaciens enzymologie   genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• fosfotransferasy s alkoholovou skupinou jako akceptorem * MeSH
• fosfotransferasy MeSH
• hydroxymočovina MeSH
• rekombinantní DNA MeSH
• ribonukleotidreduktasy MeSH
• streptomycin 3''-kinase MeSH Prohlížeč

Alizarin glucosyl transferase activity was found in five mutant strains of Streptomyces aureofaciens. The activity bears no direct relationship to the final products of tetracycline biosynthesis.

• MeSH
• anthrachinony farmakologie   MeSH
• glukosidy metabolismus   MeSH
• glukosyltransferasy izolace a purifikace   metabolismus   MeSH
• mutageneze MeSH
• Streptomyces aureofaciens enzymologie   genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• alizarin MeSH Prohlížeč
• anthrachinony MeSH
• glukosidy MeSH
• glukosyltransferasy MeSH

Kinetic parameters kcat and KM were measured for cleavage of poly I, poly A, poly U, poly C and poly I poly C by guanyl-specific RNases Sa, Pb1 and T1 and compared with that of guanyl-preferential RNase Bi. Catalytic efficiencies of the investigated enzymes to polynucleotide substrates vary considerably. The structural basis for specificity of these RNases is discussed. A hypothesis is suggested that Ser-56 plays an important role in recognition of poly A by RNase Bi.

• MeSH
• Aspergillus oryzae enzymologie   MeSH
• endoribonukleasy metabolismus   MeSH
• guanyloribonukleasa metabolismus   MeSH
• kinetika MeSH
• Penicillium enzymologie   MeSH
• polynukleotidy metabolismus   MeSH
• ribonukleasa P MeSH
• ribonukleasy metabolismus   MeSH
• RNA katalytická metabolismus   MeSH
• Streptomyces aureofaciens enzymologie   MeSH
• substrátová specifita MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• endoribonukleasy MeSH
• guanyloribonukleasa MeSH
• polynukleotidy MeSH
• ribonuclease S MeSH Prohlížeč
• ribonukleasa P MeSH
• ribonukleasy MeSH
• RNA katalytická MeSH

The crystal structures of ribonuclease from Streptomyces aureofaciens (RNase Sa) and its complex with 3'-guanylic acid (guanosine 3'-monophosphate, 3'-GMP) have been determined by the method of isomorphous replacement. The atomic parameters have been refined by restrained least-squares minimization using data in the resolution range 10.0-1.8 A. All protein atoms and more than 230 water atoms in the two crystal structures have been refined to crystallographic R factors of 0.172 and 0.175 respectively. The estimated r.m.s. error in the atomic positions ranges from 0.2 A for well-defined atoms to about 0.5 A for more poorly defined atoms. There are two enzyme molecules in the asymmetric unit, built independently, and referred to as molecules A and B. The value of the average B factor for protein atoms in both structures is about 19 A2 and for water molecules about 35 A2. Electron density for the substrate analogue 3'-GMP was found only at the active site of molecule A. The density was very clear and the positions of all 3'-GMP atoms were refined with precision comparable to that of the protein.

• MeSH
• chemické jevy MeSH
• chemie fyzikální MeSH
• difrakce rentgenového záření MeSH
• konformace proteinů MeSH
• krystalizace MeSH
• kyselina 5'-guanylová metabolismus   MeSH
• molekulární sekvence - údaje MeSH
• molekulární struktura MeSH
• ribonukleasy chemie   metabolismus   MeSH
• sekvence aminokyselin MeSH
• Streptomyces aureofaciens enzymologie   MeSH
• vazebná místa MeSH
• vodíková vazba MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• 3'-guanylic acid MeSH Prohlížeč
• kyselina 5'-guanylová MeSH
• ribonukleasy MeSH

An RNA polymerase-binding 167 bp HinfI fragment from a low-copy Streptomyces plasmid pSA 2201 has been shown to have promoter activity in vivo using a promoter-probe vector. This promoter (A1) is probably involved in expression of the genes responsible for the production of an antibiotic compound, found to be located on this plasmid. A 2600 nucleotides (nt) long transcript starting from this promoter has been identified by Northern hybridization analysis. The transcription start point has been determined using high-resolution S1 mapping and confirmed by in vitro transcription analysis with purified S. aureofaciens 2201 RNA polymerase. The A1 promoter shows no homology in the -10 and -35 consensus sequence of the typical bacterial promoters, and expression from this promoter is temporally dependent on the phase of growth having the maximum transcription activity in the stationary phase.

• MeSH
• autoradiografie MeSH
• bakteriální geny MeSH
• DNA řízené RNA-polymerasy genetika   MeSH
• elektroforéza v polyakrylamidovém gelu MeSH
• genetická transkripce MeSH
• hybridizace nukleových kyselin MeSH
• molekulární sekvence - údaje MeSH
• northern blotting MeSH
• plazmidy MeSH
• promotorové oblasti (genetika) * MeSH
• regulace genové exprese u bakterií MeSH
• sekvence aminokyselin MeSH
• sekvenční homologie nukleových kyselin MeSH
• Streptomyces aureofaciens enzymologie   genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• DNA řízené RNA-polymerasy MeSH

By chromatography on phosphocellulose and Heparin-Sepharose the modification methylase M.Sau3239I was detected and partly purified from cells of Streptomyces aureofaciens 3239. Methylation by this enzyme protects DNA from cleavage by the restriction endonuclease R.Sau3239I. The enzyme catalyzes methylation of adenine to N-6-methyladenine in the 5'-CTCGmAG-3' recognition sequence.

• MeSH
• cytosin-specifické DNA-methylasy izolace a purifikace   MeSH
• denaturace nukleových kyselin MeSH
• DNA metabolismus   MeSH
• metylace MeSH
• restrikční endonukleasy typu II MeSH
• Streptomyces aureofaciens enzymologie   genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• CTCGAG-specific type II deoxyribonucleases MeSH Prohlížeč
• cytosin-specifické DNA-methylasy MeSH
• DNA MeSH
• restrikční endonukleasy typu II MeSH