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J N Hobbs

Publications and source records attributed to J N Hobbs.

16 recordsLinked to original sources
PubMed1997

Molecular interactions of a semisynthetic glycopeptide antibiotic with D-alanyl-D-alanine and D-alanyl-D-lactate residues.

LY191145 is an N-alkylated glycopeptide antibiotic (the p-chlorobenzyl derivative of LY264826) with activity against vancomycin-susceptible and -resistant bacteria. Similar to vancomycin, LY191145 inhibited polymerization of peptidoglycan when muramyl pentapeptide served as a substrate but not when muramyl tetrapeptide was used, signifying a substrate-dependent mechanism of inhibition. Examination of ligand binding affinities for LY191145 and the effects of this agent on R39 D,D-carboxypeptidase action showed that, similar to vancomycin, LY191145 had an 800-fold greater affinity for N,N'-diacetyl-L-Lys-D-Ala-D-Ala than for N,N'-diacetyl-L-Lys-D-Ala-D-Lac. The antibacterial activity of LY191145 was antagonized by N,N'-diacetyl-L-Lys-D-Ala-D-Ala, but the molar excess required for complete suppression exceeded that needed to suppress inhibition by vancomycin. LY191145 is strongly dimerized and the p-chlorobenzyl side chain facilitates interactions with bacterial membranes. These findings are consistent with a mechanism of inhibition where interactions between antibiotic and D-Ala-D-Ala or D-Ala-D-Lac residues depend on intramolecular effects occurring at the subcellular target site.

Anti-Bacterial Agents↗
PubMed1997

The role of hydrophobic side chains as determinants of antibacterial activity of semisynthetic glycopeptide antibiotics.

Vancomycin, LY264826 and four N-substituted derivatives of LY264826 were examined for dimerization, binding to D-alanyl-D-alanine- and D-alanyl-D-lactate-containing cell wall ligands, and binding to bacterial membrane vesicles. The six glycopeptide antibiotics represent a 360-fold range in antibacterial activities against Micrococcus luteus (MIC = 0.00072-0.26 microM) with the N-substituted compounds having the lowest MICs. Vancomycin, LY264826 and the four N-substituted derivatives shared nearly identical binding affinities for N,N'-diacetyl-L-lysyl-D-alanyl-D-alanine (Kb = 1.5 x 10(5) approximately 5.9 x 10(5) M-1). Affinities for binding N,N'-diacetyl-L-lysyl-D-alanyl-D-lactate were lower but also represented a narrow range (Kb = 0.24 x 10(3) approximately 1.6 x 10(3) M-1). In contrast to ligand binding, the relative capacity of the six compounds to dimerize differed by four orders of magnitude (Kdim = 4.9 x 10(1)-1.2 x 10(6) M-1). The N-substituted derivatives had the highest Kdim values, required the greatest molar excess of exogenous cell wall ligand to suppress inhibition, and demonstrated a propensity to bind to bacterial membrane vesicles. The derivatives with the most lipophilic side chains were the most highly bound to vesicles. The findings suggest that the enhanced antibacterial activities of N-substituted derivatives of LY264826 derive from the nature of the hydrophobic side chain which can have a marked effect on dimerization and membrane binding.

Anti-Bacterial Agents↗
PubMed1996

Inhibition of peptidoglycan biosynthesis in vancomycin-susceptible and -resistant bacteria by a semisynthetic glycopeptide antibiotic.

LY191145 is a p-chlorobenzyl derivative of LY264826 (A82846B) with activity against both vancomycin-susceptible and -resistant enterococci. Incorporation of L-[14C]lysine into peptidoglycan of intact vancomycin-susceptible and -resistant Enterococcus faecium was inhibited by LY191145 (50% inhibitory concentrations of 1 and 5 microgram/ml, respectively). Inhibition was accompanied by accumulation of UDP-muramyl-peptide precursors in the cytoplasm. This agent inhibited late-stage steps in peptidoglycan biosynthesis in permeabilized E. faecium when either the UDP-muramyl-pentapeptide precursor from vancomycin-susceptible E. faecium or the UDP-muramyl-pentadepsipeptide precursor from vancomycin-resistant E. faecium was used as a substrate. Inhibition of late-stage steps led to accumulation of an N-acetyl-[14C]glucosamine-labeled lipid intermediate indicative of inhibition of the transglycosylation step. Inhibition of peptidoglycan polymerization without affecting cross-linking in a particulate membrane-plus-wall-fragment assay from Aerococcus viridans was consistent with this explanation. The fact that inhibition of peptidoglycan biosynthesis by LY191145 was not readily antagonized by an excess of free acyl-D-alanyl-D-alanine or acyl-D-alanyl-D-lactate ligands indicates that the manner in which this compound inhibits transglycosylation may not be identical to that of vancomycin.

Anti-Bacterial Agents↗
PubMed1996

Knockout of the two ldh genes has a major impact on peptidoglycan precursor synthesis in Lactobacillus plantarum.

Most bacteria synthesize muramyl-pentapeptide peptidoglycan precursors ending with a D-alanyl residue (e.g., UDP-N-acetylmuramyl-L-Ala-gamma-D-Glu-L-Lys-D-Ala-D-Ala). However, it was recently demonstrated that other types of precursors, notably D-lactate-ending molecules, could be synthesized by several lactic acid bacteria. This particular feature leads to vancomycin resistance. Vancomycin is a glycopeptide antibiotic that blocks cell wall synthesis by the formation of a complex with the extremity of peptidoglycan precursors. Substitution of the terminal D-alanine by D-lactate reduces the affinity of the antibiotic for its target. Lactobacillus plantarum is a lactic acid bacterium naturally resistant to vancomycin. It converts most of the glycolytic pyruvate to L- and D-lactate by using stereospecific enzymes designated L- and D-lactate dehydrogenases, respectively. In the present study, we show that L. plantarum actually synthesizes D-lactate-ending peptidoglycan precursors. We also report the construction of a strain which is deficient for both D- and L-lactate dehydrogenase activities and which produces only trace amounts of D- and L-lactate. As a consequence, the peptidoglycan synthesis pathway is drastically affected. The wild-type precursor is still present, but a new type of D-alanine-ending precursor is also synthesized in large quantities, which results in a highly enhanced sensitivity to vancomycin.

Anti-Bacterial Agents↗
PubMed1995

Induction of vancomycin resistance in Enterococcus faecium by non-glycopeptide antibiotics.

Bacitracin and other antibiotics that inhibit late stages in peptidoglycan biosynthesis induce vancomycin resistance in a high-level, inducibly vancomycin-resistant strain of Enterococcus faecium. Exposure to bacitracin led to synthesis of the lactate-containing UDP-MurNAc-pentadepsipeptide precursor required for vancomycin resistance. These findings indicate that inhibition of peptidoglycan biosynthesis can lead to induction of vancomycin resistance and raise the possibility that multiple signals may serve to induce resistance.

Anti-Bacterial Agents↗
PubMed1992

Biosynthesis of modified peptidoglycan precursors by vancomycin-resistant Enterococcus faecium.

In the presence of bacitracin, vancomycin-resistant Enterococcus faecium (vanA phenotype) accumulate UDP-N-acetylmuramyl(UDP-Mur-NAc)-tetrapeptide and a UDP-MurNAc-depsipentapeptide containing lactate substituted for the carboxy-terminal-D-alanine residue. In an in vitro peptidoglycan polymerization assay, the modified precursors function and confer resistance to vancomycin.

Amino Acid Sequence↗
PubMed1991

Inhibition of membrane potential-dependent amino acid transport by daptomycin.

Daptomycin inhibits the formation of UDP-N-acetylmuramyl-pentapeptide in Bacillus megaterium by inhibiting active transport of amino acids incorporated into the pentapeptide. The ability of daptomycin to inhibit active transport and peptidoglycan formation may be due to its ability to disrupt the transmembrane electrochemical gradient.

Alanine↗
PubMed1990

Inhibition of peptidoglycan biosynthesis in Bacillus megaterium by daptomycin.

The effects of daptomycin on exponential phase cells of Bacillus megaterium were investigated. Bacteriostasis was observed for concentrations between 1 and 3 micrograms/ml and maximal rate of cell lysis at 10 micrograms/ml. At sublytic concentrations (1.5-3 micrograms/ml), the variations of the pools of UDP-N-acetylglucosamine and UDP-N-acetylmuramyl-pentapeptide, as well as the incorporation of (14C)-N-acetylglucosamine into peptidoglycan were studied. From the results it was concluded that the lethal target of daptomycin could be a metabolic step between glucosamine 6-phosphate and UDP-N-acetylglucosamine.

Acetylglucosamine↗
PubMed1990

A54145 a new lipopeptide antibiotic complex: microbiological evaluation.

A54145 complex is made up of eight factors; A, A1, B, B1, C, D, E, and F which were active in vitro (MIC 0.25 approximately greater than 32 micrograms/ml) against Gram-positive aerobic organisms. The complex, factor B and B1 were found to be active against two strains of Clostridium perfringens. A calcium dependence study on some of the factors showed that their in vitro antibacterial activity was greatly enhanced by the presence of calcium (50 mg/liter) in the media. Resistance build-up was seen when Staphylococcus sp. and Streptococcus sp. were passed seven times in the presence of sublethal concentrations of A54145 antibiotics. This resistance disappeared immediately when the resistant organisms were passed in the absence of the antibiotics. Factor A was very effective against Staphylococcus aureus and Streptococcus pyogenes infections in mice (sc ED50s of 3.3 approximately 2.4 mg/kg x 2, respectively). Factor B was more active against S. pyogenes in vivo (sc ED50, 0.9 mg/kg x 2). Acute mouse toxicities were determined with these antibiotics. Semisynthetic derivatives were evaluated.

Amino Acid Sequence↗
PubMed1990

Antibacterial properties of the bicyclic pyrazolidinones.

LY173013 and LY186826 are bicyclic pyrazolidinones containing a novel aza-gamma-lactam ring structure. The antibacterial properties of these compounds appear to be related to those of beta-lactam antibiotics in that both classes of compounds share certain common binding molecules such as beta-lactamases and penicillin-binding proteins.

Alanine↗
PubMed1987

Inhibition of peptidoglycan biosynthesis in gram-positive bacteria by LY146032.

LY146032, a cyclic lipopeptide antibiotic, is an inhibitor of cell wall peptidoglycan biosynthesis in gram-positive bacteria. Although LY146032 at relatively high concentrations inhibited the in vitro polymerization of UDP-linked sugar precursors, inhibition of cell wall formation in intact Staphylococcus aureus and Bacillus megaterium cells did not lead to the accumulation of UDP-N-acetyl-muramyl (MurNAc)-peptide(s). Experiments that measured formation of UDP-MurNAc-peptides revealed that LY146032 inhibited the formation of these nucleotide-linked intermediates. This antibiotic had a disruptive effect on membrane permeability as evidenced by the loss of intracellular potassium immediately after exposure to the drug. The lack of any major disruption of the phosphoenolpyruvate:sugar phosphotransferase system indicated that the membrane is not likely a lethal target for this antibiotic. The findings are consistent with a mechanism by which LY146032 inhibits the formation of precursor molecules utilized in peptidoglycan biosynthesis. The observed membrane effects likely result from transit of the inhibitor to its lethal target site.

Anti-Bacterial Agents↗
PubMed1984

Expression of prokaryotic genes for hygromycin B and G418 resistance as dominant-selection markers in mouse L cells.

Novel bacterial resistance genes were cloned and expressed as dominant selection markers in mammalian cells. Escherichia coli genes coding for resistance to the aminocyclitol antibiotics hygromycin B (Hm) and G418 were cloned into the eukaryotic expression plasmid pSV5GPT [Mulligan and Berg, Proc. Natl. Acad. Sci. USA 78 (1981) 2072-2076]. Mouse cells normally sensitive to 100 micrograms/ml Hm were transformed with these plasmids and selected in 200 micrograms/ml Hm. Transformants resistant to as much as 1 mg/ml Hm and 500 micrograms/ml G418 were isolated. Cell extracts contained an acetyltransferase activity capable of acetylating G418 and an Hm aminocyclitol phosphotransferase activity. Plasmid DNA sequences were identified by Southern blot analysis of high Mr DNA isolated from transformed cells.

Acetyltransferases↗
PubMed1983

Genetic and enzymatic basis of hygromycin B resistance in Escherichia coli.

A plasmid conferring resistance to the aminocyclitol antibiotic hygromycin B was isolated from Escherichia coli. The gene conferring resistance to this drug was cloned in pBR322, and the gene was localized to a fragment of ca. 1,510 base pairs. Resistance to hygromycin B is determined by an aminocyclitol phosphotransferase that modifies hygromycin B and structurally related antibiotics. The specific modification of hygromycin B is a phosphorylation of the hydroxyl on the 4 position of the cyclitol ring (hyosamine). The presence of the phosphotransferase in E. coli correlates with reduced accumulation of [14C]hygromycin B.

Acetyltransferases↗
PubMed1982

7-Hydroxytropolone: an inhibitor of aminoglycoside-2"-O-adenylyltransferase.

Aminoglycoside-2"-O-adenylyltransferase was inhibited by 7-hydroxytropolone. Inhibition was competitive with respect to the cosubstrate ATP and appeared to require the unique vicinal arrangement of oxygens found in 7-hydroxytropolone. Combinations of 7-hydroxytropolone plus the appropriate aminoglycoside substrates were active against resistant bacteria possessing the adenylyltransferase. No potentiation was observed against other aminoglycoside-resistant or -susceptible strains. The fact that the inhibition of an aminoglycoside-modifying enzyme overcomes the poor uptake of aminoglycosides in resistant strains points to the singular importance of the inactivating enzyme as a determinant of resistance.

Aminoglycosides↗