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G R Drapeau

Publications and source records attributed to G R Drapeau.

At least 19 recordsLinked to original sources
PubMed1992

The rcsB gene, a positive regulator of colanic acid biosynthesis in Escherichia coli, is also an activator of ftsZ expression.

Wild-type genes which, when overexpressed, are capable of restoring the growth deficiency of the division mutant ftsZ84 of Escherichia coli on L medium containing no added NaCl have been isolated. One of these genes is rcsB, a positive regulator of colanic acid biosynthesis. A direct relationship between rcsB expression and FtsZ activity was observed, suggesting that RcsB specifically increases transcription of ftsZ, thus accounting for the restoration of colony formation by ftsZ84 mutant cells. Analysis of the 5' upstream sequence of rcsB revealed, in addition to the sigma 54 promoter sequence previously reported, a presumptive sigma 70 promoter and LexA-binding site plus an upstream sequence that is found to be essential for the expression of rcsB on a plasmid. The absence of the sigma 54 factor does not have a negative effect on the transcription of rcsB. The RcsB protein is an activator of its own synthesis, particularly in the presence of NaCl. Evidence which suggests that RcsB can be phosphorylated by a presumably modified EnvZ or PhoM sensor protein leading to a suppression of the growth deficiency of ftsZ84 mutant cells and to an increase in colanic acid production was obtained. We also demonstrated that the level of colanic acid is reduced when the cells carry a multicopy rcsC plasmid, suggesting that the RcsC sensor has phosphatase activity.

Amino Acid Sequence↗
PubMed1992

Identification, cloning, and characterization of rcsF, a new regulator gene for exopolysaccharide synthesis that suppresses the division mutation ftsZ84 in Escherichia coli K-12.

A new gene, designated rcsF, was located adjacent to drpA at the 5.2-min position of the genetic map of Escherichia coli. The deduced amino acid sequence encoded by the rcsF gene indicates a small protein of 133 amino acid residues with a calculated pI of 10.8 that is rich in proline, serine, alanine, and cysteine residues. When overexpressed as a result of its presence on a multicopy plasmid, rcsF confers a mucoid phenotype and restores colony formation to ftsZ84 mutant cells on L agar medium containing no added NaCl. These two phenotypes are not observed in rcsB mutant cells. Ion mutant cells harboring an rcsF mutation accumulate considerably lower levels of exopolysaccharides, whereas the presence of a multicopy rcsF plasmid not only increases capsule synthesis but also confers a mucoid phenotype at 37 degrees C, a temperature at which ion mutant cells are known not to form mucoid colonies. RcsF does not stimulate the expression of rcsB, indicating that it exerts its action through the RcsB protein, possibly by phosphorylation. It is also shown that RcsF stimulation of capsule synthesis is RcsA-dependent, whereas colony formation of ftsZ84 mutant cells can be restored by RcsF in the absence of RcsA.

Amino Acid Sequence↗
PubMed1990

Mutations conferring resistance to azide in Escherichia coli occur primarily in the secA gene.

Mutant strains of Escherichia coli were screened for the ability to grow on L agar plates containing 3.4 or 4.6 mM sodium azide. Most mutants had mutations located in the leucine region, presumably at the azi locus. Two of these mutants were found to have a mutation in the secA gene, but expression of the resistance phenotype also required the presence of upstream gene X. While a plasmid carrying the X-secA mutant gene pair was able to confer azide resistance to a sensitive host, a similar plasmid harboring the wild-type secA allele rendered a resistant strain sensitive to azide, indicating codominance of the two alleles. That azide inhibits SecA is consistent with the fact that SecA has ATPase activity, an activity that is often prone to inhibition by azide.

Adenosine Triphosphatases↗
PubMed1990

Molecular cloning, nucleotide sequence, and expression of shl, a new gene in the 2-minute region of the genetic map of Escherichia coli.

Cells of Escherichia coli that harbor supH (an allele of the wild-type gene serU) are sensitive to UV irradiation and temperature and appear to have an impaired cell division control mechanism. We found that a gene located at the 2-min region, designated shl, inhibited the growth of supH-harboring cells when carried by a high-copy-number plasmid, whereas the same plasmid had no visible effect when present in parental cells. The amino acid sequence predicted from the nucleotide sequence of the shl gene indicated a similarity to the GalR and LacI repressor proteins, suggesting it is a transcription regulator. The sequence between the promoter and the structural genes revealed the presence of a short open reading frame of 28 amino acid residues followed by a segment of 81 base pairs. These structural features suggest that a transcription antitermination mechanism may be involved in the regulation of expression of the shl gene. The possibility that shl is a regulator of serU is discussed.

Amino Acid Sequence↗
PubMed1989

The Escherichia coli cell division mutation ftsM1 is in serU.

The ftsM1 mutation is believed to be in a gene implicated in the regulation of cell division in Escherichia coli because it displayed the lon mutation phenotypes. In this study, we show that this mutation is located in serU, a gene which codes for tRNA(Ser)2, and has the phenotypes of the serU allele supH. Both ftsM1 and supH suppressed the leuB6 and ilvD145 missense mutations, and both conferred temperature and UV light irradiation sensitivity to the harboring cells. Cells which carried the ftsM1 mutation or the supH suppressor had very low colony-forming abilities on salt-free L agar, and this phenotype was almost completely abolished by the presence of plasmids bearing the ftsZ+ gene. Furthermore, sensitivity of the mutant cells to UV irradiation was also markedly diminished when they carried a ftsZ+-bearing plasmid. These results suggest that supH-containing cells have reduced FtsZ activities, in accordance with their displaying the phenotypes of the lon mutant cells. The possibility that ftsM1 (supH) is functionally involved in the biosynthesis of a specific protein which affects cell division is discussed.

Alleles↗
PubMed1988

Cell division control in Escherichia coli K-12: some properties of the ftsZ84 mutation and suppression of this mutation by the product of a newly identified gene.

The Fts proteins play an important role in the control of cell division in Escherichia coli. These proteins, which possibly form a functional complex, are encoded by genes that form an operon. In this study, we examined the properties of the temperature-sensitive mutation ftsZ84 harbored by low- or high-copy-number plasmids. Cells of strain AB1157, which had the ftsZ84 mutation, did not form colonies on salt-free L agar at 30 degrees C. When a low-copy-number plasmid containing the ftsZ84 mutation was present in these mutant cells, colony formation was restored on this medium at 30 degrees C, suggesting that FtsZ84 is probably less active than the wild-type protein and is therefore limiting in its capacity to trigger cell divisions. On the other hand, when the ftsZ84 mutation was harbored by the high-copy-number plasmid pBR325, colony formation was prevented on salt-free L agar plates whether the recipients were ftsZ84 mutant or parental cells, suggesting that, at high levels, FtsZ84 acts as a division inhibitor. The fact that colony formation was also prevented at 42 degrees C indicates that the FtsZ84 protein is not inactivated at the nonpermissive temperature. The possibility that FtsZ84 is a more efficient division inhibitor than the wild-type FtsZ is discussed. Evidence is also presented showing that a gene adjacent to mutT codes for a product that, under certain conditions, suppresses the ftsZ84 mutation.

Bacterial Proteins↗
PubMed1987

Regulation of cell division in Escherichia coli K-12: probable interactions among proteins FtsQ, FtsA, and FtsZ.

In Escherichia coli, the FtsQ, FtsA, and FtsZ proteins are believed to play essential roles in the regulation of cell division. Of the three proteins, FtsZ has received the most attention, particularly because of its interactions with SfiA. Double mutants which carry mutations located in the ftsQ, ftsA, or ftsZ gene in combination with the lon-1 mutation were constructed. In the presence of the lon-1 mutation, which is known to stabilize SfiA, the ftsQ1 mutant cells were not capable of forming colonies on a rich agar medium, whereas mutant cells harboring either one of the mutations grew well on this medium. Examination of lon-1 fts double-mutant cells for sensitivity to UV light revealed that those carrying the ftsA10 allele were resistant. It was also observed that in the presence of a multicopy plasmid containing a wild-type ftsZ gene, the ftsQ1 mutant filamented markedly following a nutritional shift-up and that the division rate of ftsZ84 mutant cells was slightly reduced when they harbored a wild-type ftsQ-containing plasmid. The possibility that the Fts proteins are interacting with one another and forming a molecular complex is discussed.

ATP-Dependent Proteases↗
PubMed1986

Identification of new cell division genes in Escherichia coli by using extragenic suppressors.

To facilitate the analysis of the cell division control apparatus in Escherichia coli, we studied extragenic suppressor mutations of a previously characterized temperature-sensitive division mutation, ftsM1. Cells of strain GD40 which harbor this mutation were spread on agar plates and incubated at 42 degrees C, and the surviving cells were analyzed for the presence of a suppressor mutation. One group of suppressed mutants had acquired a new mutation which, by conjugation, was found to be located in the 30- to 40-min region of the E. coli genetic map. The other group comprised revertants carrying a suppressor which appeared to map between thr and leu. This suppressor gene, called sftA, was cloned with a mini-Mu-derived in vivo cloning system by selection for suppression of temperature sensitivity in GD40 cells. Subsequent subcloning of a fragment of the chromosomal DNA from the mini-Mu plasmid into pBR325 resulted in the delineation of the suppressor gene on a 1.8-kilobase XhoI-PvuI fragment. A strain, CV514, which does not express the temperature sensitivity phenotype of the ftsM1 mutation, was found to harbor a natural suppressor of this mutation. UV sensitivity, another known phenotype of the ftsM1 mutation, was also corrected by the presence of the sftA suppressor in the cell. Thus, the characterization of extragenic suppressors may allow the identification of new genes involved in the control of cell division.

Cell Division↗
PubMed1984

Regulation of cell division in Escherichia coli: properties of new ftsZ mutants.

Cells of Escherichia coli which produce high levels of the sfiA protein are UV-sensitive and filament extensively. It has been postulated that the sfiA protein is a division inhibitor which interacts with the ftsZ protein (formerly called sfiB or sulB) leading to cell division arrest. Under certain conditions, a similar division inhibition is observed with cells harboring a mutationally altered tsM allele, another division gene which was postulated to code for a division inhibitor or a controlling effector thereof (Drapeau et al. (1984). In this communication, we report on the properties of ftsZ mutants isolated under conditions which brought no selective pressure. These mutants have either an increased sensitivity to UV irradiation or filament drastically following a nutritional shift-up, or both, or even cannot grow in a rich medium. They presumably possess a ftsZ protein which responds more readily to the inhibitory action of the wild type sfiA or the mutationally altered tsM1 protein since the phenotypic expressions associated with the mutations are not observed in the presence of the sfiA11 mutation or are amplified when the ftsZ mutant cells harbor the tsM1 allele. These results further support earlier suggestions that sfiA modulates ftsZ activity and establish tsM as an additional regulatory element thereof. In addition, it is shown that E. coli strain B is a naturally occurring ftsZ mutant.

Bacterial Proteins↗
PubMed1984

Regulation and SOS induction of division inhibition in Escherichia coli K12.

When Escherichia coli is subjected to treatments that damage DNA or perturb DNA replication considerable cell filamentation occurs. It has been postulated that this phenomenon is associated with the presence of a division inhibitor induced coordinately with the SOS functions. The role of this induction would be to delay septation during DNA repair to prevent the formation of DNAless cells. In this communication, we present evidence for such a division inhibitor based on the properties of a division mutant which is hyperactive in the septation delay. Cells of this mutant filament extensively after a nutritional shift-up, have drastically reduced colony-forming abilities on a rich medium but not on a minimal medium following treatment with ultraviolet radiation and, are deficient in the lysogenization of phage lambda; phenotypes which are characteristic of but expressed to a much lower extent in another type of division mutant called Ion. Cells harboring the division mutation plus either one of the lexA mutant alleles, spr-51 or tsl-1, are filamentous suggesting that they are permanently derepressed for division inhibition. These results are in agreement with models that assign the regulation of cell division to a division inhibitor which is regulated by the lexA repressor protein.

Bacteriophage lambda↗
PubMed1983

Unusual properties of a new division mutant of Escherichia coli.

The properties of a division mutant of Escherichia coli were investigated. At 42 degrees C, this mutant forms nonseptate, multinucleate, filamentous cells typical of division mutants, and at the permissive temperature, is sensitive to ultraviolet (UV) irradiation. Temperature and UV sensitivities are probably due to a single mutation. The mutant phenotype is dominant to wild type. The mutant cells make DNA nearly as effectively as control cells at 42 degrees C or following UV irradiation. They exhibit normal host-cell reactivation capacities and can express all manifestations of the SOS response with the exception of Weigle reactivation. The genetic lesion which mediates this pleiotropic effect is located very close to the leu locus of the linkage map.

Bacteriophage lambda↗
PubMed1980

Substrate specificity of a proteolytic enzyme isolated from a mutant of Pseudomonas fragi.

Previous studies have described the isolation of mutationally altered proteases in Pseudomonas fragi (Noreau, J., and Drapeau, G.R. (1979) J. Bacteriol, 140, 911-916. In the present study, it is shown that one of these proteases cleaves specifically the peptide bonds on the NH2-terminal side of either aspartic acid or cysteic acid residues in oxidized ribonuclease. With myoglobin as the substrate, a similar specificity was observed except that only four out of the six aspartyl bonds present were hydrolyzed.

Amino Acids↗
PubMed1980

Prothrombin activation by a metalloprotease from Staphylococcus aureus.

Formation of thrombin during incubation of purified bovine prothrombin with purified staphylococcal metalloprotease has been investigated. Thrombin activity was estimated by examination of clotting time and by digestion of a synthetic substrate, Chromozym TH. The metalloprotease caused direct activation of prothrombin which was inhibited by the addition of ethylenediaminetetraacetic acid. Metalloprotease produced by some strains of Staphylococcus aureus may simulate staphylocoagulase activity.

Animals↗
PubMed1979

Isolation and properties of the protease from the wild-type and mutant strains of Pseudomonas fragi.

A simplified procedure for the purification of the extracellular protease of Pseudomonas fragi was developed. The enzyme was isolated from a derepressed mutant producing 40 times the enzyme level of the parental organism. It was collected from culture filtrates by ammonium sulfate precipitation, and it was obtained in pure form by single chromatography on a column of diethylaminoethyl cellulose. The protease had a molecular weight of 52,000 as estimated by sodium dodecyl sulfate-gel electrophoresis and had properties of a classical neutral endopeptidase with the exception of its substrate specificity. Mutants of P. fragi producing proteases of altered substrate specificities were isolated from plates containing elastin as the sole carbon source. The SP-Sephadex elution patterns of enzymes extracted from each mutant examined were complex, suggesting that either the enzyme was autodigested or several active forms could be generated from a common precursor. The substrate specificities of the mutant enzymes were different from that produced by the parental strain.

Amino Acids↗
PubMed1978

Unusual COOH-terminal structure of staphylococcal protease.

The extracellular enzyme, staphylococcal protease, carries a COOH-terminal tryptic peptide of 43 amino acid residues most of which are aspartic acid, asparagine, and proline. This peptide might have a function equivalent to that of a similar segment previously observed at the NH2-terminal end of the membrane-bound penicillinase precursor of Bacillus licheniformis (Yamamoto, S., and Lampen, J. O. (1976) Proc. Natl. Acad. Sci. U. S. A. 73, 1457-1461). These observations would suggest that bacterial exoproteins which are secreted in the form of precursors differ from extracellular proteins by the presence of an extra segment at their NH2- and/or COOH-terminal ends.

Amino Acid Sequence↗
PubMed1978

Role of metalloprotease in activation of the precursor of staphylococcal protease.

A metalloprotease was isolated from the culture medium of a mutant of Staphylococcus aureus strain V8. The enzyme had a molecular weight of 38,000 as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and an optimum pH of 7.0 and exhibited a specificity for peptide bonds on the N-terminal side of large hydrophobic residues. The protease was fully inactivated by 0-phenanthroline but could be reactivated by zinc ions. Cobalt may be substituted for zinc, producing an activity which corresponds to 160% of that of the native enzyme. All these data indicate that this protease is a typical bacterial neutral metalloprotease. The role of this metalloprotease in the activation of the precursor of another protease secreted by the same organism, staphylococcal protease, has been identified. Mutants which lack the metalloprotease accumulated the precursor, which can be specifically activated by the addition of the purified metalloprotease or the related enzyme thermolysin. The purification of the precursor is also reported.

Amino Acids↗
PubMed1978

The primary structure of staphylococcal protease.

The amino acid sequence of staphylococcal protease has been determined by analysis of tryptic peptides obtained from cyanogen bromide fragments. Selected peptides obtained from digests with staphylococcal protease, thermolysin, and chymotrypsin provided the information necessary to align the tryptic peptides and the cyanogen bromide fragments. The protease is a single polypeptide chain of some 250 amino acids and is devoid of sulfhydryl groups. The COOH-terminal tryptic peptide of of the protease molecule contains some 43 residues, most of which are aspartic acids, asparagines, and prolines. The amino acid sequence of this peptide was not determined. The primary structure near the active serine residue indicates that staphylococcal protease is related to the pancreatic serine proteases. However, it has little or no additional sequence homologies with these enzymes except for the regions near histidine-50 and aspartic acid - 91. These regions have striking similarities with the corresponding regions of protease B and the trypsin-like enzyme of Streptomyces griseus.

Amino Acid Sequence↗