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J Houmard

Publications and source records attributed to J Houmard.

43 records · Page 3Linked to original sources
PubMed1980

The use of translocatable genetic elements to construct a fine-structure map of the Klebsiella pneumoniae nitrogen fixation (nif) gene cluster.

The transposons Tn5, Tn7 and Tn10 and bacteriophage Mu have been used to derive insertion mutations in the Klebsiella pneumoniae nif gene cluster. A large number of deletion mutants have been derived by imprecise excision of insertion mutations and these deletions have been used to construct a fine-structure map of the nif cluster. Comparison of this genetic map with a physical map of the nif cluster derived by Reidel et al. (1979) showed a very good correlation between genetic and physical mapping methods. A new complementation group, designated nifU, has been identified and mapped between nifN and nifS. Polarity studies on the 14 nif cistrons now identified suggests that they are organized in at least seven transcriptional units and that all the multicistronic units are transcribed in the same direction.

Chromosome Deletion↗
PubMed1978

Genetic and biochemical analysis of mutants induced by bacteriophage Mu DNA integration into Klebsiella pneumoniae nitrogen fixation genes.

Polar mutations were obtained by integration of bacteriophage Mu c+ or Mu cts DNA into the Klebsiella pneumoniae nif genes located on plasmid pCE1, a derivative of pRD1. In addition, nif deletions were isolated from nif::Mu cts plasmids. Complementation data allowed the characterization of twelve nif cistrons, nine corresponding to previously identified genes. Polar effect of Mu DNA insertions suggested the existence of at least six transcription units: 1) nif K, nif D and nif H--2)nif A and nif L--3) nif E and a new gene--4) nif B--5) nif F--6) nif J. Nif K, nif D and nif H, which are most probably the structural genes for nitrogenase, seem to belong to the same operon transcribed from nif H to nif K. This was confirmed by SDS gel autoradiography of pulse labelled proteins. Moreover it was possible to identify, on the autoradiograms, a polypeptide which likely is the product of nif J and whose biosynthesis is under the control of nif A.

Coliphages↗
PubMed1976

Kinetic investigation of the staphylococcal protease-catalyzed hydrolysis of synthetic substrates.

In investigating the staphylococcal protease-catalyzed hydrolysis of N-tert-butoxycarbonyl-L-glutamate alpha-phenyl ester, N-benzyloxycarbonyl-L-glutamate alpha-phenyl ester and N-benzyloxycarbonyl-L-glutamate alpha-p-nitroanilide, we obtained kinetic evidence consistent with the formation of an acyl-enzyme intermediate. We found that addition of a nucleophile, such as methanol, led to the partition of the common acyl-enzyme intermediate between water and the alcohol. With N-benzyl-oxycarbonyl-L-glutamate alpha-phenyl ester, a specific ester substrate, deacylation was shown to be the rate-limiting step. By studying the kcat/Km ratio of these hydrolyses as a function of pH, we have shown that two ionizable groups on the enzyme are essential to the catalytic process. One of these groups has a pK of 6.58 and the other, a pK of 8.25. The assignment of these pK values is discussed in connection with the known features of the serine proteinase reaction mechanism. In addition, monovalent anions were shown to inhibit staphylococcal protease hydrolyses. They seem to compete with the negative charge of the substrate, thus inhibiting its binding on the enzyme molecule. Finally we compared the kinetic parameters obtained with five proteases isolated from different strains of Staphylococcus aureus.

Buffers↗
PubMed1976

Preparation of chromophoric substrates for the glutamoyl specific staphylococcal protease.

The synthesis of chromophoric substrates allowing an accurate determination of the staphylococcal protease activity is described. BOC-L-Glu-OPh, BOC-L-Phe-L-Glu-OPh, BOC-L-Ala-L-Glu-OPh, BOC-L-Ser-L-Glu-OPh and Z-L-Glu-pNA were prepared. Kinetic parameters of the staphyloccal protease-catalysed hydrolyses of these substrates are compared. In every case the dipeptide ester substrates lead to a lower catalytic efficiency (kcat/Km ratio), compared with either BOC-L-Glu-OPh or Z-L-Glu-OPh, mainly because of an increase in the Km value. Like other serine proteinases, the staphylococcal protease exhibits a high ratio of eeterase to peptidase activity, the kcat/Km ratio being 2.6 X 10(5)-fold higher with the Z-L-Glu-OPh than with the Z-L-Glu-pNA.

Anilides↗
PubMed1972

Staphylococcal protease: a proteolytic enzyme specific for glutamoyl bonds.

An extracellular protease of Staphylococcus aureus, strain V8, previously shown to cleave specifically the peptide bonds on the carboxyl-terminal side of either aspartate or glutamate residues in phosphate buffer (pH 7.8) hydrolyzes only glutamoyl bonds in either ammonium bicarbonate (pH 7.8) or ammonium acetate (pH 4.0). Of all aspartoyl bonds tested, only the Asp-Gly linkage is cleaved at a detectable rate. The staphylococcal protease hydrolyzes all of the seventeen different glutamoyl bonds studied, although those involving hydrophobic aminoacid residues with bulky side chains are cleaved at a lower rate.

Amino Acid Sequence↗