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M Mevarech

Publications and source records attributed to M Mevarech.

35 records · Page 2Linked to original sources
PubMed1988

Crystallization of halophilic malate dehydrogenase from Halobacterium marismortui.

Malate dehydrogenase from the extreme halophile Halobacterium marismortui crystallizes in highly concentrated phosphate solution in space group 12 with cell dimensions a = 113.8 A, b = 122.8 A, c = 126.7 A, beta = 98.1 degrees. The halophilic enzyme was found to be unstable at lower concentrations of phosphate. It associates with unusually large amounts of water and salt, and the combined particle volume shows a tight fit in the unit cell.

Halobacterium↗
PubMed1988

Cloning and comparative sequence analysis of the gene coding for isopenicillin N synthase in Streptomyces.

The genes coding for isopenicillin N synthase (IPNS) in Streptomyces jumonjinensis and S. lipmanii were isolated from recombinant phage lambda libraries using the S. clavuligerus IPNS gene as a heterologous probe. The S. jumonjinensis IPNS gene has an open reading frame coding for 329 amino acids, identical in size to that of the previously cloned S. clavuligerus IPNS gene. A partial nucleotide sequence was also determined for the S. lipmanii IPNS gene. Comparison of the predicted amino acid sequences of all three streptomycete IPNS proteins shows that they exhibit more than 70% similarity, close to that found in comparisons among fungal IPNS proteins and significantly greater than that found, approximately 60%, between Streptomyces and fungal IPNS proteins. We conclude that procaryotic and eucaryotic IPNS genes are subgroups of a single family of microbial IPNS genes. Hybridization probes prepared from IPNS genes of the above streptomycete species were used to detect analogous genes in eight other strains that included both penicillin and cephalosporin producers and non-producers. Each producer strain responded with all three probes implying the presence of an IPNS gene. Surprisingly, several non-producer strains also responded with one or two of the probes. Our results suggest that IPNS-related genes may be more prevalent in Streptomyces than previously believed.

Amino Acid Sequence↗
PubMed1988

Cloning and nucleotide sequence determination of the isopenicillin N synthetase gene from Streptomyces clavuligerus.

The isopenicillin N synthetase (IPNS) gene from Streptomyces clavuligerus was isolated from an Escherichia coli plasmid library of S. clavuligerus genomic DNA fragments using a 44-mer mixed oligodeoxynucleotide probe. The nucleotide sequence of a 3-kb region of the cloned fragment from the plasmid, pBL1, was determined and analysis of the sequence showed an open reading frame that could encode a protein of 329 amino acids with an Mr of 36,917. When the S. clavuligerus DNA from pBL1 was introduced into an IPNS-deficient mutant of S. clavuligerus on the Streptomyces vector pIJ941, the recombinant plasmid was able to complement the mutation and restore IPNS activity. The protein coding region of the S. clavuligerus IPNS gene shows about 63% and 62% similarity to the Cephalosporium acremonium and Penicillium chrysogenum IPNS nucleotide sequences, respectively, and the predicted amino acid sequence of the encoded protein showed about 56% similarity to both fungal sequences.

Acremonium↗
PubMed1985

Isolation of genes from Candida albicans by complementation in Saccharomyces cerevisiae.

A genomic library of the asexual pathogenic yeast Candida albicans was constructed in the S. cerevisiae vector YEp13. The library contains a representation of the entire genome with a probability of 99%. The expression of the genes of C. albicans in S. cerevisiae was examined and two mutations his3-1 and trp1-289 of S. cerevisiae were complemented by the cloned genes of C. albicans. The hybridization data indicates that the plasmids complementing the mutations of S. cerevisiae contain sequences from C. albicans.

Candida albicans↗
PubMed1985

Genetic transfer in Halobacterium volcanii.

Auxotrophic mutants of Halobacterium volcanii generated by chemical mutagenesis were used to demonstrate a native genetic transfer system in this extremely halophilic member of the class Archaeobacteria.

Crosses, Genetic↗
PubMed1984

Regulatory region of the metA gene of Escherichia coli K-12.

Transcription of the metA gene of Escherichia coli K-12 is from a promoter which is under methionine control and is located next to a region which has an extensive sequence homology with the operator regions of the metBL and metF genes. However, in the metA gene there is a second transcription start point which is located 74 base pairs upstream and which is independent of the intracellular methionine concentration.

Acyltransferases↗
PubMed1984

RNA polymerase activity in virions from Ustilago maydis.

RNA polymerase activity is associated with the double-stranded RNA virions of Ustilago maydis. The reaction products of the polymerase activity are single-stranded RNA molecules. The RNA molecules synthesized are homologous to the three classes of double-stranded RNA molecules that typify the viral genome. The single-stranded RNA synthesized is released from the virions. The molecular weight of the single-stranded RNA transcripts is about half the size of the double-stranded RNA segments, and thus, it appears that in the in vitro reaction, full-length transcripts can be obtained.

Basidiomycota↗
PubMed1980

Amino acid sequence of 2Fe-2S ferredoxin from an extreme halophile, Halobacterium of the Dead Sea.

The primary structure of the 2Fe-2S ferredoxin from Halobacterium of the Dead Sea was determined and it consisted of 128 amino acid residues including an N epsilon-acetyllysyl residue. Due to a high degree of sequence homology between this ferredoxin and the one from Halobacterium halobium, all tryptic peptides could be aligned in order. Only 20 amino acid differences were observed between these two halobacterial ferredoxins. The distribution of cysteinyl residues involved in the iron chelation was similar to that of chloroplast-type ferredoxins.

Amino Acid Sequence↗
PubMed1980

Nucleotide sequence of a cyanobacterial nifH gene coding for nitrogenase reductase.

The nucleotide sequence of nifH, the structural gene for nitrogenase reductase (component II or Fe protein of nitrogenase) from the cyanobacterium Anabaena 7120 has been determined. Also reported are 194 bases of the 5'-flanking sequence and 170 bases of the 3'-flanking sequence. The predicted amino acid sequence was compared with that determined for the complete nitrogenase reductase of Clostridium pasteurianum and the cysteine-containing peptides of the protein from Azotobacter vinelandii. Amino acid sequences around five cysteines, located in the NH(2)-terminal two-thirds of the protein, are highly conserved in all three species. Codon usage in the first gene from a cyanobacterium to be sequenced shows striking asymmetries for eight amino acids.

Journal Article↗
PubMed1979

Detection of gastrin-specific mRNA using oligodeoxynucleotide probes of defined sequence.

Three oligodeoxynucleotides have been chemically synthesized and used as hybridization probes to detect gastrin-specific mRNA within an heterogeneous population of RNA molecules. Using these oligonucleotides whose nucleotide sequences were deduced from the amino acid sequence of the hormone, 0.2 fmol of gastrin mRNA can be detected/microgram of poly(A)-RNA from hog antrums. The 32P-labeled oligonucleotides hybridize specifically to RNA covalently coupled to DBM-paper (Alwine, J.C., Kemp, D.J., and Stark, G.R. (1977) Proc. Natl. Acad. Sci. U.S.A. 74, 5350-5354). Labeled oligonucleotides also hybridize specifically to RNA separated by gel electrophoresis in the presence of CH3HgOH and covalently coupled to DBM-paper. Using this procedure, we have found that the mRNA coding for gastrin contains about 620 nucleotides, which is in agreement with results obtained using gastrin cDNA to determine mRNA size.

Animals↗
PubMed1979

Detection and partial sequence analysis of gastrin mRNA by using an oligodeoxynucleotide probe.

We have used a specific deoxyoligonucleotide probe to detect gastrin mRNA in poly(A)-enriched RNA preparations from hog antrum. The nucleotide sequence of the oligonucleotide, d(C-T-C-C-T-C-C-A-T-C-C-A), was deduced from the unique amino acid sequence Trp-Met-Glu-Glu of gastrin. When used with hog antral RNA, the dodecanucleotide is an effective primer for the synthesis of gastrin-specific cDNA as judged by nucleotide sequence analysis of cDNA isolated by polyacrylamide gel electrophoresis. We have determined an 81-nucleotide sequence corresponding to the region of the gastrin mRNA that codes for the known amino acid sequence of the G34 progastrin intermediate species, and we have demonstrated the presence of two consecutive basic residues preceding the G34 sequence in the prohormone. Hybridization of gastrin cDNA or synthetic dodecanucleotide to hog antral RNA separated by gel electrophoresis on agarose gels in the presence of methylmercuric hydroxide indicates that the mRNA coding for gastrin is about 620 nucleotides long. These results suggest that the gastrin precursor peptide contains 110-140 amino acids. This method should be of general application for detection and characterization of mRNAs corresponding to proteins of known amino acid sequence.

Animals↗
PubMed1977

Malate dehydrogenase isolated from extremely halophilic bacteria of the Dead Sea. 1. Purification and molecular characterization.

The complete purification of malate dehydrogenase (EC 1.1.1.37) from extremely halophilic bacteria of the Dead Sea is described. The purification procedure includes (a) precipitation by ammonium sulfate, (b) fractionation on Sepharose 4B using a decreasing concentration gradient of ammonium sulfate, (c) gel permeation chromatography on Sephadex G-100, (d) chromatography on hydroxylapatite, and (e) affinity chromatography on 8-(6-aminohexyl)amino-NAD+-Sepharose at 4.26 M NaCl. The absorption and fluorescence spectra of the native and denatured enzyme were measured, and the extinction coefficient at 280 nm in 4.26 M NaCl was found to be 0.803 cm2mg-1. The amino acid composition analysis showed an excess of 10.4 mol % of acidic amino acids. The apparent specific "volume" phi' of the active enzyme at 4.26 M NaCl was found to be 0.680 +/- 0.015 mL/g. The molecular weight of the native enzyme was found to be 84 000 +/- 4000 determined in 4.26 M NaCl from equilibrium sedimentation data. The molecular weight of the subunits is 39 000 as measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Thus, the native enzyme is composed of two subunits.

Amino Acids↗
PubMed1976

Hydrophobic chromatography and fractionation of enzymes from extremely halophilic bacteria using decreasing concentration gradients of ammonium sulfate.

Ammonium sulfate fractionation of proteins from extremely halophilic bacteria on Sepharose 4B, carboxymethylcellulose, diethylaminoethylcellulose, and hexamethylenediamine-Agarose is described. Halophilic proteins are absorbed on these gels at 2.5 M ammonium sulfate and eluted by decreasing concentration gradients of this salt. The method has enabled the separation of malate dehydrogenase from glutamate dehydrogenase and aspartate aminotransferase on Sepharose 4B and the additional 15-fold purification of glutamate dehydrogenase on DEAE-cellulose. The technique is simple and convenient, operates at low cost, and possesses great power of resolution. The mechanism of adsorption is discussed and compared to previous instances of "hydrophobic chromatography". It is concluded that the retention of halophilic proteins on the polysaccharide gels at 2.5 M ammonium sulfate is due to hydrophobic interactions.

Ammonium Sulfate↗