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Journal of Bacteriology Aug 1965Pruess, David L. (University of Wisconsin, Madison), and Marvin J. Johnson. Enzymatic deacylation of S(35)-benzylpenicillin. J. Bacteriol. 90:380-383....
Pruess, David L. (University of Wisconsin, Madison), and Marvin J. Johnson. Enzymatic deacylation of S(35)-benzylpenicillin. J. Bacteriol. 90:380-383. 1965.-S(35)-benzylpenicillin, penicilloic acid, and penilloic acid were deacylated by cell suspensions of Escherichia coli and Micrococcus roseus. Both cultures deacylated penicillin most rapidly and penilloic acid least rapidly. The deacylase activity of M. roseus against penicilloic acid was cell-bound, probably requiring a metal ion for activity.
Topics: Acetone; Acylation; Buffers; Centrifugation; Chemical Phenomena; Chemistry; Chromatography; Culture Media; Escherichia coli; Fermentation; Hydrogen-Ion Concentration; Metabolism; Micrococcus; Penicillanic Acid; Penicillin G; Penicillins; Pharmacology; Phenylacetates; Radiometry; Research; Sulfur Isotopes
PubMed: 14329451
DOI: 10.1128/jb.90.2.380-383.1965 -
European Journal of Biochemistry Sep 1980Streptococcus faecalis ATCC 9790 possesses seven membrane-bound penicillin-binding proteins. They have been characterized with respect to their apparent molecular...
Streptococcus faecalis ATCC 9790 possesses seven membrane-bound penicillin-binding proteins. They have been characterized with respect to their apparent molecular weights, relative abundance, specificity profiles for 15 different beta-lactam antibiotics and stability under various conditions. In water and at 37 degrees C, all the native penicillin-binding proteins have half-lives longer than 20 h except protein 3b (half-life of about 600 min) and protein 4 (half-life of about 175 min). The short-lived 80 000-Mr protein 4 is spontaneously converted into a 73 000-Mr water-soluble, penicillin-binding protein 4. Similarly, the short-lived 82 000-Mr protein 3b seems to be the protein from which the 72 000-Mr water-soluble protein X spontaneously originates during incubation of the membranes. Release of both proteins 4 and X from the membrane is maximal under alkaline conditions; it is not inhibited by various protease inhibitors. After exposure to trypsin, the 43 000-Mr membrane-bound penicillin binding protein 6 (a DD-carboxypeptidase) gives to a 30 000-Mr water-soluble protein 6. Like the parent protein, protein 6 exhibits both DD-carboxypeptidase activity and penicillin-binding ability. With proteins 6 and 6, low dose levels of p-chloromercuribenzoate prevent both enzyme activity and combination with penicillin, thus strongly suggesting that a thiol group is involved in the enzyme active center. We have shown previously [Coyette et al. in Eur. J. Biochem. 88, 297--305 (1978) and 75, 231--239 (1977)] that the DD-carboxypeptidase protein 6 fragments the benzylpenicillin molecule with formation of phenylacetylglycine. Breakdown of the complex formed between [14C]benzylpenicillin and 14 000-Mr membrane-bound protein 1 is also 'enzyme-catalysed'. Most likely, however, the released product is penicilloate. With all the other penicillin-binding proteins whose molecular weights are intermediate between those of proteins 1 and 6, breakdown of the complexes formed with [14C]benzylpenicillin results from proteolysis and is not due to the release of the bound metabolite. None of the penicillin-binding proteins behaves, by itself, as a lethal target for beta-lactam antibiotic action on the living cells.
Topics: Binding, Competitive; Carrier Proteins; Cell Membrane; Enterococcus faecalis; Half-Life; Kinetics; Molecular Weight; Muramoylpentapeptide Carboxypeptidase; Penicillin G; Penicillins
PubMed: 6777159
DOI: 10.1111/j.1432-1033.1980.tb04886.x -
PloS One 2015Antibiotics, such as benzyl-penicillin (PenG) and cephalosporin, are the most common compounds used in animal therapy. Their massive and illegal use in animal therapy...
Antibiotics, such as benzyl-penicillin (PenG) and cephalosporin, are the most common compounds used in animal therapy. Their massive and illegal use in animal therapy and prophylaxis inevitably causes the presence of traces in foods of animal origin (milk and meat), which creates several problems for human health. With the aim to prevent the negative impact of β-lactam and, in particular, PenG residues present in the milk on customer health, many countries have established maximum residue limits (MRLs). To cope with this problem here, we propose an effective alternative, compared to the analytical methods actually employed, to quantify the presence of penicillin G using the surface plasmon resonance (SPR) method. In particular, the PenG molecule was conjugated to a protein carrier to immunize a rabbit and produce polyclonal antibodies (anti-PenG). The produced antibodies were used as molecular recognition elements for the design of a competitive immune-assay for the detection of PenG by SPR experiments. The detection limit of the developed assay was found to be 8.0 pM, a value much lower than the MRL of the EU regulation limit that is fixed at 12 nM. Thus, our results clearly show that this system could be successfully suitable for the accurate and easy determination of PenG.
Topics: Animals; Anti-Bacterial Agents; Antibodies; Chromatography, Affinity; Drug Residues; Enzyme-Linked Immunosorbent Assay; Limit of Detection; Milk; Penicillin G; Surface Plasmon Resonance
PubMed: 26168259
DOI: 10.1371/journal.pone.0132396 -
Applied Microbiology Jan 1963Benzylpenicillin acylase, which hydrolyzes benzylpenicillin to 6-aminopenicillanic acid, was found to be widely distributed among members of the Schizomycetes,...
Benzylpenicillin acylase, which hydrolyzes benzylpenicillin to 6-aminopenicillanic acid, was found to be widely distributed among members of the Schizomycetes, particularly in gram-negative bacteria, and in the genus Nocardia. The hydrolysis of a series of biosynthetic and semisynthetic penicillins by freeze-dried cells of a strain of Nocardia and of Proteus was studied. Benzylpenicillin was the preferred substrate; all departures from the benzylpenicillin side-chain structure led to reduction of substrate activity (the greater the departure, the greater the reduction in activity). Penicillin amides and methyl esters were also hydrolyzed, as were suitable N-acyl derivatives of 7-aminocephalosporanic acid. Occurrence of an enzyme activity which hydrolyzes benzylpenicillinamide to benzylpenicillin was detected in certain strains of yeasts.
Topics: Cephalosporins; Esterases; Nocardia; Penicillanic Acid; Penicillin Amidase; Penicillin G; Penicillins; Proteus; Substrate Specificity
PubMed: 13955341
DOI: 10.1128/am.11.1.1-6.1963 -
The Biochemical Journal Dec 19691. A method is given for the preparation of penicillin acylase by using Escherichia coli N.C.I.B. 8743 and a strain selected for higher yield. The enzyme is associated...
1. A method is given for the preparation of penicillin acylase by using Escherichia coli N.C.I.B. 8743 and a strain selected for higher yield. The enzyme is associated with the bacterial cells and removes the side chains of penicillins to give 6-amino-penicillanic acid and a carboxylic acid. 2. The rates of penicillin deacylation indicated that p-hydroxybenzylpenicillin was the best substrate, followed in diminishing order by benzyl-, dl-alpha-hydroxybenzyl-, 2-furylmethyl-, 2-thienylmethyl-, d-alpha-aminobenzyl-, n-propoxymethyl- and isobutoxymethyl-penicillin. Phenylpenicillin and dl-alpha-carboxybenzylpenicillin were not substrates and phenoxymethyl-penicillin was very poor. 3. Amides and esters of the above penicillins were also substrates for the deacylation reaction, as were cephalosporins with a thienylmethyl side chain. 4. For the deacylation of 2-furylmethylpenicillin at 21 degrees the optimum pH was 8.2. The optimum temperature was 60 degrees at pH7. 5. By using selection A of N.C.I.B. 8743 and determining reaction velocities by assaying yields of 6-amino-penicillanic acid in a 10min. reaction at 50 degrees and pH8.2, the K(m) for benzylpenicillin was found to be about 30mm and the K(m) for 2-furylmethylpenicillin, about 10mm. The V(max.) values were 0.6 and 0.24mumole/min./mg. of bacterial cells respectively.
Topics: Amides; Amidohydrolases; Ampicillin; Cephalosporins; Escherichia coli; Esters; Hydrogen-Ion Concentration; Kinetics; Methicillin; Penicillin G; Penicillin V; Penicillins; Temperature
PubMed: 4982417
DOI: 10.1042/bj1150733 -
Antimicrobial Agents and Chemotherapy Jan 1980Penicillin-binding proteins of three Streptomyces strains, S. cacaoi, S. olivaceus, and S. clavuligerus, were examined by gel electrophoresis and fluorography. In a...
Penicillin-binding proteins of three Streptomyces strains, S. cacaoi, S. olivaceus, and S. clavuligerus, were examined by gel electrophoresis and fluorography. In a beta-lactamase producer, S. cacaoi, at least five membrane-bound penicillin-binding proteins were detected, but in two beta-lactam producers, S. olivaceus and S. clavuligerus, fewer penicillin-binding proteins were detected. Mecillinam and methicillin bound selectively to some penicillin-binding proteins in S. cacaoi, whereas they did not bind at all to those in S. olivaceus and S. clavuligerus. Clavulanic acid bound to penicillin-binding proteins only at a very high concentration in S. cacoi. This compound did not bind to those of S. olivaceus and S. clavuligerus. Penicillin-binding proteins in culture supernatant and cytoplasm and minimum inhibitory concentrations of these strains against benzylpenicillin were also examined.
Topics: Amdinocillin; Membrane Proteins; Methicillin; Penicillin G; Penicillin Resistance; Protein Binding; Streptomyces; beta-Lactamases
PubMed: 6965440
DOI: 10.1128/AAC.17.1.1 -
Canadian Medical Association Journal May 1982
Topics: Adolescent; Ampicillin; Drug Therapy, Combination; Female; Gonorrhea; Humans; Newfoundland and Labrador; Penicillin G
PubMed: 7074442
DOI: No ID Found -
The Biochemical Journal Mar 19651. Three variants of staphylococcal exopenicillinase (types A, B and C) can be distinguished on chemical, enzymological and immunological grounds. 2. Enzyme type A has a...
1. Three variants of staphylococcal exopenicillinase (types A, B and C) can be distinguished on chemical, enzymological and immunological grounds. 2. Enzyme type A has a higher specific activity than that of type B, but has a similar combination affinity with anti-(exopenicillinase type A) serum. 3. Enzyme types A and C have a similar specific activity, but enzyme type C has a lower combination affinity for anti-(exopenicillinase type A) serum than has enzyme type A. 4. The sedimentation coefficients and amino acid analyses of the three enzyme types are similar. 5. All three enzyme types have small but significant differences in kinetics of action when hydrolysing benzylpenicillin, methicillin, cloxacillin and cephalosporin C. 6. Peptide maps, obtained from enzyme types A and C after digestion with trypsin, show that these two variants probably differ in the nature of only a very few amino acid residues. 7. Enzyme type B seems to be confined to staphylococci that are members of staphylococcal phage group II. Enzyme types A and C are produced by staphylococci that are members either of phage group I or III, but never group II. 8. The low specific enzyme activity and affinity of enzyme type B towards all penicillins tested suggest that this enzyme type has a lower ;efficiency' in hydrolysing penicillin and therefore in protecting bacteria from the action of penicillin. This could account for the low incidence among ;hospital staphylococci' of penicillin-resistant staphylococci that are members of phage group II.
Topics: Amino Acids; Biochemical Phenomena; Biochemistry; Cephalosporins; Cloxacillin; Drug Resistance, Microbial; Immune Sera; Methicillin; Mutation; Penicillin G; Penicillin Resistance; Penicillinase; Penicillins; Peptides; Research; Staphylococcal Infections; Staphylococcus; Staphylococcus Phages; Staphylococcus aureus
PubMed: 14342258
DOI: 10.1042/bj0940584 -
Journal of Dairy Science Jul 2014The degradation of penicillin G, penicillin V, and ampicillin in milk in the presence of β-lactamase was investigated by ultra-performance liquid chromatography coupled...
The degradation of penicillin G, penicillin V, and ampicillin in milk in the presence of β-lactamase was investigated by ultra-performance liquid chromatography coupled with electrospray ionization-time-of-flight mass spectrometry. Degradation products of the 3 penicillins in milk were identified based on the fact that the metabolites or degradation products contain a substructure of penicillin, and their degradation pathways in acidic milk in presence of β-lactamase were developed. The influence of factors on the degradation was investigated, including β-lactamase dosage, temperature, time, and acidity. The ratio of the 2 degradation products (penicilloic acid and penilloic acid) is different at different temperatures and pH. Penicilloic acid was the dominant species obtained at pH 6 under 40°C, but, being unstable, it could not be used as a standard for accurate analysis of penicilloic acid, and also could not be used as target for detection of penicillins in milk. Penilloic acid was the dominant species obtained at pH 2 above 40°C; it was stable and could be used as a standard for quantitative analysis and as target for detecting whether penicillins were used in milk.
Topics: Ampicillin; Animals; Chromatography, High Pressure Liquid; Dose-Response Relationship, Drug; Hydrogen-Ion Concentration; Milk; Penicillin G; Penicillin V; Spectrometry, Mass, Electrospray Ionization; Temperature; Time Factors; beta-Lactamases
PubMed: 24792795
DOI: 10.3168/jds.2014-7952 -
Journal of Bacteriology Jul 1987Murein synthesized in ether-permeabilized cells of Escherichia coli deficient in individual penicillin-binding proteins (PBPs) and in the presence of certain beta-lactam...
Murein synthesized in ether-permeabilized cells of Escherichia coli deficient in individual penicillin-binding proteins (PBPs) and in the presence of certain beta-lactam antibiotics was analyzed by high-pressure liquid chromatography separation of the muramidase split products. PBP 1b was found to to be the major murein synthesizing activity that was poorly compensated for by PBP 1a. A PBP 2 mutant as well as mecillinam-inhibited cells showed increased activity in the formation of oligomeric muropeptides as well as UDP-muramylpeptidyl-linked muropeptides, the reaction products of transpeptidation, bypassing the lipid intermediate. In contrast, penicillin G and furazlocillin severely inhibited these reactions but stimulated normal dimer production. It is concluded that two distinct transpeptidases exist in E. coli: one, highly sensitive to penicillin G and furazlocillin, catalyzes the formation of hyper-cross-linked muropeptides, and a second one, quite resistant to these antibiotics, synthesizes muropeptide dimers.
Topics: Amdinocillin; Azlocillin; Bacterial Proteins; Carrier Proteins; Cell Division; Escherichia coli; Hexosyltransferases; Imidazolidines; Muramoylpentapeptide Carboxypeptidase; Penicillin-Binding Proteins; Penicillins; Peptidoglycan; Peptidyl Transferases
PubMed: 3298212
DOI: 10.1128/jb.169.7.3099-3103.1987