Modulation of different forms of glutamine synthetases inRhizobium phaseoli and their possible role in nitrogen fixation

The effect of a number of compounds structurally related to glutamic acid and other nitrogenous compounds on the composition of three forms of glutamine synthetase (GS) inRhizobium phaseoli has been examined in detail. Amino acids like glutamic acid, glutamine, and a fixed source of nitrogen like ammonium chloride did not alter the relative glutamine synthetase composition.l-Methioninedl-sulfoximine (MSX), a glutamate analogue, significantly repressed the synthesis of GSIII to a greater extent.,N-oxalyl,-diaminopropionic acid (ODAP), another glutamate analogue, selectively stimulated the synthesis of GSII, and the effect of ODAP on GSII synthesis was greatly enhanced in the presence of ethylenediamine or ammonium chloride. Ethylenediamine itself caused a predominant synthesis of GSIII.-Cyanoalanine-grownR. phaseoli did not synthesize GSI. The synthesis of the three different glutamine synthetases can thus be differentially modulated.     

Identification and characterization of three forms of glutamine synthetase unique to Rhizobia

Glutamine synthetase (GS) activities of Rhizobia were chromatographically resolved into three distinct forms, GSI, GSII, and GSIII on DEAE cellulose, being eluted with 0.3M, 0.5M and 0.8M KCl, respectively. GSIII was the major form inR. leguminosarum andR. phaseoli. InR. meliloti, however, GSI was the major form. The three forms of GS were also distinguished on the basis of (a) rapid heat inactivation of GSII, (b) insensitivity of GSI to inhibitors, (c) marked inhibition of GSII by thymidine, and (d) inability of Zn⁺⁺ to inhibit GSIII. The three forms of GS are also distinct molecular entities and are unique to Rhizobia.     

High-level expression of the Streptomyces clavuligerus isopenicillin N synthase gene in Escherichia coli.

The pcbC gene, which encodes isopenicillin N synthase (IPNS), was subcloned from Streptomyces clavuligerus into Escherichia coli by using the pT7 series of plasmid vectors. The polymerase chain reaction was used to introduce an NdeI site at the translation initiation codon of pcbC, allowing the gene to be inserted behind an E. coli type of ribosome binding site. This construction directed high-level expression of IPNS, but the IPNS was in an inactive form in inclusion bodies. Active IPNS was recovered by solubilizing and renaturing the protein.     

Purification and characterization of recombinantStreptomyces clavuligerus isopenicillin N synthase produced inEscherichia coli

Recombinant isopenicillin N synthase fromStreptomyces clavuligerus was produced in the form of inactive inclusion bodies inEscherichia coli. These inclusion bodies were solubilized by treatment with 5 M urea under reducing conditions. Optimization of refolding conditions to recover active isopenicillin N synthase indicated that a dialysis procedure carried out at a protein concentration of about 1.0 mg ml^–1 gave maximal recovery of active isopenicillin N synthase. Solubilized isopenicillin N synthase of more than 95% purity was obtained by passing this material through a DEAE-Trisacryl ion exchange column. Expression studies conducted at different temperatures indicated that isopenicillin N synthase was produced predominantly in a soluble, active form when expression was conducted at 20°C, and accounted for about 20% of the total soluble protein. This high-level production facilitated the purification of soluble isopenicillin N synthase to near homogeneity in four steps. Characterization of the purified soluble and solubilized isopenicillin N synthase revealed that they are very similar.     

High-level expression of the Streptomyces clavuligerus isopenicillin N synthase gene in Escherichia coli.

The pcbC gene, which encodes isopenicillin N synthase (IPNS), was subcloned from Streptomyces clavuligerus into Escherichia coli by using the pT7 series of plasmid vectors. The polymerase chain reaction was used to introduce an NdeI site at the translation initiation codon of pcbC, allowing the gene to be inserted behind an E. coli type of ribosome binding site. This construction directed high-level expression of IPNS, but the IPNS was in an inactive form in inclusion bodies. Active IPNS was recovered by solubilizing and renaturing the protein.     

Draft genome sequencing and functional annotation and characterization of biofilm-producing bacterium Bacillus novalis PD1 isolated from rhizospheric soil

Antonie van Leeuwenhoek - Biofilm forming bacterium Bacillus novalis PD1 was isolated from the rhizospheric soil of a paddy field. B. novalis PD1 is a Gram-positive, facultatively anaerobic,...     

An In Silico Evaluation of Molecular Interaction Between Antimicrobial Peptide Subtilosin A of Bacillus subtilis with Virulent Proteins of Aeromonas hydrophila

International Journal of Peptide Research and Therapeutics - Subtilosin A, a cyclic peptide from Bacillus subtilis is known for its antimicrobial activity against a diverse range of bacteria....     

Introduction of histidine units using benzotriazolide activation

N^α‐Boc‐N^im‐(4‐toluenesulfonyl‐l ‐histidylbenzotriazole) enables convenient acylation of N‐, O‐, S‐, and C‐nucleophiles with no detectable racemization. We report efficient syntheses of novel histidine‐containing di‐, tri‐, and tetra‐peptides and models for the preparation of potentially biologically active histidine N‐, O‐, S‐, and C‐conjugates. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.