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See detailOn the DD-carboxypeptidase enzyme system of Streptomyces strain K15
Leyh-Bouille, Mélina; Nguyen-Distèche, Martine ULg; Ghuysen, Jean-Marie ULg

in European Journal of Biochemistry (1981), 115(3), 579-584

Streptomyces K15 possesses a set of exocellular and cell-bound D-alanyl-D-alanine carboxypeptidases. Four of them have been isolated to the stage where each enzyme preparation contains on single ... [more ▼]

Streptomyces K15 possesses a set of exocellular and cell-bound D-alanyl-D-alanine carboxypeptidases. Four of them have been isolated to the stage where each enzyme preparation contains on single penicillin-binding protein. The exocellular 54000-Mr enzyme is extremely sensitive to benzylpenicillin and performs low transpeptidase activity on the carbonyl-donor/amino-acceptor tetrapeptide ACLLys(Gly)-DAla-DAla. The exocellular 40 000-Mr enzyme and the two lysozyme-releasable 40 000-Mr and 38 000-Mr enzymes are moderately sensitive to benzylpenicillin and have a high propensity to catalyse dimer formation from the aforementioned tetrapeptide monomer. [less ▲]

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See detailLocalisation of the phosphoester bond hydrolyzed by the major apurinic/apyrimidinic endodeoxyribonuclease from rat-liver chromatin.
Verly, Walter G; Colson, Pierre ULg; Zocchi, Germaine ULg et al

in European Journal of Biochemistry (1981), 118

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See detailThe penicillin-binding proteins in Streptococcus faecalis ATCC 9790
Coyette, Jean; Ghuysen, Jean-Marie ULg; Fontana, Roberta

in European Journal of Biochemistry (1980), 110(2), 445-456

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 ... [more ▼]

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. [less ▲]

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See detailKinetic mechanism of the exchanges catalysed by the adenine-nucleotide carrier.
Duyckaerts, Claire ULg; Sluse-Goffart, Claudine; Fux, Jean-pierre et al

in European Journal of Biochemistry (1980), 106

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See detailKinetic and binding properties of the oxoglutarate translocator of rat-heart mitochondria.
Sluse, Francis ULg; Duyckaerts, Claire ULg; Sluse-Goffart, Claudine et al

in European Journal of Biochemistry (1979), 100

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See detailSolubilization and isolation of the membrane-bound DD-carboxypeptidase of Streptococcus faecalis ATCC9790. Properties of the purified enzyme
Coyette, Jacques; Ghuysen, Jean-Marie ULg; Fontana, Roberta

in European Journal of Biochemistry (1978), 88(1), 297-305

Streptococcus faecalis ATCC 9790 possesses six membrane-bound, penicillin-binding proteins. That numbered 6 (Mr 43000) is the most abundant one and is the DD-carboxypeptidase studied previously. The ... [more ▼]

Streptococcus faecalis ATCC 9790 possesses six membrane-bound, penicillin-binding proteins. That numbered 6 (Mr 43000) is the most abundant one and is the DD-carboxypeptidase studied previously. The enzyme has been solubilized and purified to the stage where one single protein band can be detected by gel electrophoresis. The purification procedure does not alter the properties that the enzyme exhibits when it is membrane-bound. The DD-carboxypeptidase itself may be a killing target for penicillin in S. faecalis. [less ▲]

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See detailInteraction between penicillin and the DD-carboxypeptidase of the unstable L-form of Proteus mirabilis strain 19
Schilf, Wolfgang; Frere, Philippe; Frère, Jean-Marie ULg et al

in European Journal of Biochemistry (1978), 85(2), 325-330

Binding of penicillin to the DD-carboxypeptidase of the unstable spheroplast L-form of Proteus mirabilis results in the rapid formation of a modified enzyme-inhibitor complex which in turn undergoes rapid ... [more ▼]

Binding of penicillin to the DD-carboxypeptidase of the unstable spheroplast L-form of Proteus mirabilis results in the rapid formation of a modified enzyme-inhibitor complex which in turn undergoes rapid decay into reactivated enzyme and an antibiotically inactive penicillin degradation product. Major antibiotic metabolites recovered from such interactions were benzylpenicilloic acid and phenoxymethylpenicilloic acid from benzylpenicillin and phenoxymethylpenicillin, respectively, suggesting a second enzymic function of the DD-carboxypeptidase as a penicillinase of low efficiency. Statistical analyses made with the help of a linear regression program show that the enzyme interacts with the substrate UDP-N-acetylmuramoyl-L-alanyl-D-gamma-glutamyl-(L)-meso-2,6-diaminopimelyl -(L)-D-alanyl-D-alanine and either benzympenicillin or carbenicillin in a non-competitive manner. [less ▲]

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See detailThe peptidoglycan crosslinking enzyme system in Streptomyces R61, K15 and rimosus. Immunological studies
Nguyen-Distèche, Martine ULg; Frère, Jean-Marie ULg; Dusart, Jean et al

in European Journal of Biochemistry (1977), 81(1), 29-32

The exocellular DD-carboxypeptidases from Streptomyces R61, K 15, the lysozyme-releasable DD-carboxypeptidases from Streptomyces R61, K15 and rimosus, and the membrane-bound DD-carboxypeptidase of ... [more ▼]

The exocellular DD-carboxypeptidases from Streptomyces R61, K 15, the lysozyme-releasable DD-carboxypeptidases from Streptomyces R61, K15 and rimosus, and the membrane-bound DD-carboxypeptidase of Streptomyces K15 are immunologically related to each other. [less ▲]

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See detailThe peptidoglycan crosslinking enzyme system in Streptomyces strains R61, K15 and rimosus : Kinetic Coefficients Involved in the Interactions of the Membrane-Bound Transpeptidase with Peptide Substrates and β-Lactam Antibiotics
Dusart, Jean; Leyh-Bouille, Mélina; Ghuysen, Jean-Marie ULg

in European Journal of Biochemistry (1977), 81(1), 33-44

The transpeptidation reaction performed by the membranes of Streptomyces strain R61 fits the general rate equation for an enzyme-catalysed bimolecular reaction. The same membranes (E) interact with beta ... [more ▼]

The transpeptidation reaction performed by the membranes of Streptomyces strain R61 fits the general rate equation for an enzyme-catalysed bimolecular reaction. The same membranes (E) interact with beta-lactams (I) to form inactive penicillin-enzyme-membrane complexes (EI) of rather high stability, which subsequently break down (E + I leads to EI leads to E + degradation products). The enzyme is regenerated and the antibiotic is released in the form of an inactive metabolite. With benzylpenicillin, the degradation product is benzylpenicilloic acid. The reaction is heat-labile. The first step of the reaction (E + I leads to EI) is characterized by a second-order rate constant (kformation in M-1 s-1) and the second step (EI leads to E + degradation products) by a first-order rate constant (kbreakdown in s-1). The effects in vitro of various beta-lactams on the membrane-bound transpeptidase, as expressed by the relevant kformation and kbreakdown values, parallel the effects in vivo of the same antibiotics as expressed by their ability to prevent the germination and growth of conidiospores. The kinetic parameters of the transpeptidase that was solubilized with N-cetyl-N,N,N-trimethylammonium bromide with respect to its interaction with both peptide substrates and beta-lactam antibiotics are quantitatively different from those of the membrane-bound enzyme. Moreover, the solubilized enzyme fragments benzylpenicillin with formation of phenylacetylglycine, a reaction which is similar to that catalysed by the exocellular R61 enzyme. The membranes of Streptomyces strains rimosus and K15 possess an active 'classic' penicillinase. They were not studied but the kinetic coefficients of the corresponding solubilized transpeptidases were determined and compared with those of the solubilized enzyme from strain R61. [less ▲]

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See detailInteractions between beta-lactam antibiotics and isolated membranes of Streptococcus faecalis ATCC 9790
Coyette, Jacques; Ghuysen, Jean-Marie ULg; Binot, Françoise ULg et al

in European Journal of Biochemistry (1977), 75(1), 231-239

The DD-carboxypeptidase-exchange membrane-bound enzyme in Streptococcus faecalis ATCC 9790 reacts with beta-lactam antibiotics to form complexes with rather long half-lives. Depending upon the antibiotic ... [more ▼]

The DD-carboxypeptidase-exchange membrane-bound enzyme in Streptococcus faecalis ATCC 9790 reacts with beta-lactam antibiotics to form complexes with rather long half-lives. Depending upon the antibiotic, the second-order rate constants for complex formation range from 0.75-560 M-1 S-1 (at 37 degrees C and in water) and the first-order rate constants for complex breakdown range from 1.3 to 26 x 10(-5) s-1 (at 37 degrees C and in 5 mM phosphate buffer pH 7.5). There are about 30 pmol of DD-carboxypeptidase-exchange enzyme per mg of membrane protein. The degradation products arising from benzylpenicillin are phenylacetylglycine and probably N-formyl-D-penicillamine. Isolated membranes also contain other penicillin binding sites (about 70 pmol/mg membrane protein). That part of benzylpenicillin which reacts with at least some of these latter sites is slowly degraded into penicilloic acid. Normal functioning of the DD-carboxypeptidase-exchange membrane-bound enzyme is important, if not essential, for cell growth. With the beta-lactam antibiotics tested inhibition of cell growth is mainly related to the rates of formation of the inactive enzyme-antibiotic complexes. The relationship, however, is not a direct one probably due to the competitive effect exerted by the other penicillin binding sites. [less ▲]

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See detailThe exchange reaction of peptides R-D-alanyl-D-alanine with D-[14C]alanine to R-D-alanyl-D-[14C]alanine and D-alanine, catalysed by the membranes of Streptococcus faecalis ATCC 9790
Coyette, Jacques; Ghuysen, Jean-Marie ULg; Perkins, Harold R.

in European Journal of Biochemistry (1977), 75(1), 225-229

Under alkaline conditions, the membrane-bound DD-carboxypeptidase of Streptococcus faecalis ATCC 9790 catalyses exchange reactions in which the X-L-R3-D-Ala moiety of peptides of the type X-L-R3-D-Ala-D ... [more ▼]

Under alkaline conditions, the membrane-bound DD-carboxypeptidase of Streptococcus faecalis ATCC 9790 catalyses exchange reactions in which the X-L-R3-D-Ala moiety of peptides of the type X-L-R3-D-Ala-D-Ala is transferred to simple amino compounds such as D-alanine, glycine and glycyl-glycine. The enzyme system is unable, however, to catalyse complex reactions that would simulate the natural transpeptidation reaction. [less ▲]

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See detailThe peptidoglycan crosslinking enzyme system in Streptomyces strains R61, K15 and rimosus. Exocellular, lysozyme-releasable and membrane-bound enzymes.
Leyh-Bouille, Mélina; Dusart, Jean; Nguyen Van, Martine ULg et al

in European Journal of Biochemistry (1977), 81(1), 19-28

The DD-carboxypeptidase (I)-transpeptidase (II) system in Streptomyces strain K15 consists of: (1) a membrane-bound II capable of performing low I activity; and (2) a set of I: (a) membrane-bound, (b ... [more ▼]

The DD-carboxypeptidase (I)-transpeptidase (II) system in Streptomyces strain K15 consists of: (1) a membrane-bound II capable of performing low I activity; and (2) a set of I: (a) membrane-bound, (b) lysozyme-releasable, and (c) exocellular, having low II activities in aq. media and at low acceptor concns. The I are related to each other and may belong to the same pathway leading to enzyme excretion. A similar system occurs in Streptomyces strain R61 except that the membrane-bound I activity is low when compared with the membrane-bound II activity. In S. rimosus, the system consists almost exclusively of the membrane-bound II and the levels of membrane-bound, lysozyme-releasable, and exocellular I are very low. [on SciFinder(R)] [less ▲]

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See detailActivity and Subcellular Distribution of Protein Kinase Dependent on Adenosine 3': 5'-Monophosphate in Liver from Normal and Adrenalectomized Rats
Rousseau, Guy; Martial, Joseph ULg; De Visscher, M.

in European Journal of Biochemistry (1976), 66(3), 499-506

We have examined whether glucocorticoids control the activity and (or) the subcellular distribution of protein kinase dependent on cyclic AMP (adenosine 3':5'-monophosphate), since they influence cyclic ... [more ▼]

We have examined whether glucocorticoids control the activity and (or) the subcellular distribution of protein kinase dependent on cyclic AMP (adenosine 3':5'-monophosphate), since they influence cyclic-AMP-dependent responses to other hormones. Protein kinase activity was determined in rat liver homogenates and subcellular fractions, nuclear, large granular, microsomal and supernatant obtained by differential sedimentation in 0.25 M sucrose. 63% of the tissue protein kinase activity detected in absence of cyclic AMP reside in the particulate fractions. Upon addition of exogenous cyclic AMP, protein kinase activity is stimulated 1.8, 1.2, 1.2 and 4.5-fold in nuclear, large granular, microsomal and supernatant fractions, respectively. Under these conditions, 66% of tissue activity are found in the supernatant fraction. The activity sensitive to exogenous cyclic AMP resolves into a major (84%) cytosoluble and a minor (16%) nucleomicrosomal component. The latter activity resists elution with isotonic saline and is increased in the presence of Triton X-100. Three groups of rats were studied: control and adrenalectomized with or without cortisol treatment. In whole liver homogenates, both protein kinase activity detected in absence of exogenous cyclic AMP and sensitivity of the enzyme to cyclic AMP were comparable in all groups. Moreover, the distribution patterns of proteins kinase activity amoung the fractions were essentially the same in all groups of animals, whether or not particles had been treated with Triton X-100. Finally, in cell-free experiments, glucocorticoids alone or in combination with their intracellular receptor did not modify protein kinase activity of rat liver. Thus the results reported do not support the possibility that glucocorticoids influence cyclic AMP-dependent protein kinase in rat liver. Yet, this study provides data, not available before, on subcellular distribution of this enzyme in rat liver. [less ▲]

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See detailInteraction between the exocellular DD-carboxypeptidase-transpeptidase from Streptomyces R61, substrate and beta-lactam antibiotics. A choice of models
Frère, Jean-Marie ULg; Ghuysen, Jean-Marie ULg; Perkins, Harold R

in European Journal of Biochemistry (1975), 57(2), 353-359

The interaction between the exocellular DD-carboxypeptidase-transpeptidase of Streptomyces R61 and beta-lactam antibiotics is a multistep process during which a rather stable enzyme - antibiotic complex ... [more ▼]

The interaction between the exocellular DD-carboxypeptidase-transpeptidase of Streptomyces R61 and beta-lactam antibiotics is a multistep process during which a rather stable enzyme - antibiotic complex is formed. This mechanism of interaction is compatible with Lineweaver-Burk plots that are typical of a competitive inhibition of the hydrolysis of the peptide donor by the antibiotic. In fact, however, the same Lineweaver-Burk plots can be obtained on the basis of a non-competitive type of inhibition. At present, a choice between the two models cannot be made. [less ▲]

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See detailKinetics of interaction between the exocellular DD-carboxypeptidase-transpeptidase from Streptomyces R61 and beta-lactam antibiotics. A choice of models
Frère, Jean-Marie ULg; Ghuysen, Jean-Marie ULg; Iwatsubo, Motohiro

in European Journal of Biochemistry (1975), 57(2), 343-351

The simplest model for the interaction between the exocellular DD-carboxypeptidase-transpeptidase from Streptomyces R61 and beta-lactam antibiotics involves the three following steps: (a) the formation of ... [more ▼]

The simplest model for the interaction between the exocellular DD-carboxypeptidase-transpeptidase from Streptomyces R61 and beta-lactam antibiotics involves the three following steps: (a) the formation of a reversible equimolar enzyme - antibiotic complex; (b) the irreversible transformation of this complex into a modified enzyme - antibiotic complex; and (c) the breakdown of this latter complex and the concomitant release of a regenerated enzyme and a modified antibiotic molecule. The dissociation constant for step 1 and the rate constants for steps 2 and 3 were measured with various beta-lactam antibiotics. With antibiotic such as benzylpenicillin, which behaves as a good 'substrate', steps 1 and 2 occur at enzymic velocities, whereas step 3 occurs at a very low velocity and hence is responsible for the low efficiency of the overall process. [less ▲]

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See detailThe catalytic activity and penicillin sensitivity in the liquid and frozen states of membrane-bound and detergent-solubilised transpeptidase of Streptomyces R61
Dusart, Jean; Marquet, Alberto; Ghuysen, Jean-Marie ULg et al

in European Journal of Biochemistry (1975), 56(1), 57-65

The Km, app. values of the membrane-bound transpeptidase of Streptomyces R61 for the donor Ac2-L-Lys-D-Ala-D-Ala and the acceptor Gly-Gly are not affected by temperature variations when the reaction ... [more ▼]

The Km, app. values of the membrane-bound transpeptidase of Streptomyces R61 for the donor Ac2-L-Lys-D-Ala-D-Ala and the acceptor Gly-Gly are not affected by temperature variations when the reaction mixtures are incubated in liquid suspensions. At -5 degrees C, the incubation can be carried out either in the liquid or in the frozen state. The enzyme is active in the latter state. In the frozen state, the Km, app. value for the acceptor remains unchanged but there is a 3-fold increase in the maximum velocity, a 10-fold decrease of the Km, app. value for the donor and a 10-fold increase of the benzylpenicillin concentration required to inhibit the enzyme activity by 50% (ID50 value). Temperatures of -35 degrees C or below are required to completely inhibit the membrane-bound enzyme in the frozen state. Cetyltrimethylammonium bromide extracts the transpeptidase both from the isolated membranes and, with a much higher yield, from the intact mycelium. The extracted enzyme is not active in the frozen state, requires detergent for activity, has decreased Km, app. values for both donor and acceptor, exhibits the same sensitivity to benzylpenicillin and cephalosporin C as the membrane-bound transpeptidase (in liquid suspensions) and, like this latter enzyme, has no DD-carboxypeptidase activity. The detergent-extracted transpeptidase penetrates gels of Sephadex-100 and is not sedimented at 200 000 X g. [less ▲]

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See detailEvidence for cooperative effects in the exchange reaction catalysed by the oxoglutarate translocator of rat-heart mitochondria.
Sluse, Francis ULg; Sluse-Goffart, Claudine; Duyckaerts, Claire ULg et al

in European Journal of Biochemistry (1975), 56(1), 1-14

The initial rates of the exchange external oxoglutarate/internal malate through the inner membrane of rat-heart mitochondria, for various concentrations of the two substrates, have been reinvestigated for ... [more ▼]

The initial rates of the exchange external oxoglutarate/internal malate through the inner membrane of rat-heart mitochondria, for various concentrations of the two substrates, have been reinvestigated for an extended range of concentrations of the external oxoglutarate. This has been made possible by use of the inhibitor-stop technique that allows 100 times smaller incubation times than the centrifugation-stop technique used previously. Under the experimental conditions the uptake of the external-labelled oxoglutarate into the mitochondrial-matrix space is mediated by the oxoglutarate translocator performing a ono-to-one exchange of the anions oxoglutarate (external) and malate (internal). Two intermediary-plateau regions are observed in the kinetic saturation curve of the translocator by the external oxoglutarate, revealing a complex rate equation which is found to be the product of two one-substrate functions. Analysing these features it is shown that the model, proposed earlier, of a "double carrier" as catalyst in a rapid-equilibrium random bi-bi mechanism, is still applicable but that several external binding sites have to be considered. As already noticed the external and the internal substrates bind to their respective sites independently of each other. Furthermore, some additional requirements imposed by the observed kinetics suggest that the exchange reaction is performed by only one translocator species made of identical interacting subunits. The anion exchange is tentatively viewed as a rotation of a subunit around an axis situated in the plane of the membrane after two independent local configuration changes induced by the binding of the two substrates on this subunit. [less ▲]

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See detailInteraction between beta-lactam antibiotics and exocellular DD-carboxypeptidase-transpeptidase of Streptomyces R61
Frère, Jean-Marie ULg; Leyh-Bouille, Mélina; Ghuysen, Jean-Marie ULg et al

in European Journal of Biochemistry (1974), 50(1), 203-214

On the basis of steady-state kinetics, inhibition of the exocellular dd-carboxypeptidase-trans-peptidase of Streptomyces R61 by β-lactam antibiotics was competitive with regard to the donor substrate ... [more ▼]

On the basis of steady-state kinetics, inhibition of the exocellular dd-carboxypeptidase-trans-peptidase of Streptomyces R61 by β-lactam antibiotics was competitive with regard to the donor substrate. However, the complexes formed between the Streptomyces R61 enzyme and various β-lactam antibiotics were relatively stable, exhibiting half-lives of 40 to 80 min at 37°C and neutral pH. During breakdown of the complexes the protein underwent reactivation, whereas the released antibiotic molecule was chemically altered. With [14C]benzylpenicillin, the released compound was neither benzylpenicillin nor benzylpenicilloic acid. The properties of the Streptomyces R61 enzyme β-lactam antibiotic complexes were compared with those of the complexes formed between the same antibiotics and either the membrane-bound transpeptidase from Streptomyces R61 or the exocellular dd-carboxypeptidase-transpeptidase of Streptomyces R39. [less ▲]

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See detailMembrane-bound transpeptidase and penicillin binding sites in Streptomyces strain R61
Marquet, Alberto; Dusart, Jean; Ghuysen, Jean-Marie ULg et al

in European Journal of Biochemistry (1974), 46(3), 515-523

High-affinity penicillin binding sites from which the antibiotic could not be removed by washings at 4°C in 0.017 M K2HPO4 or 0.05 M Tris-HCl pH 7.5, were shown to occur in the isolated membranes of ... [more ▼]

High-affinity penicillin binding sites from which the antibiotic could not be removed by washings at 4°C in 0.017 M K2HPO4 or 0.05 M Tris-HCl pH 7.5, were shown to occur in the isolated membranes of Streptomyces R61. These sites caused the attachment of 25 picomoles of [14C]benzylpenicillin per milligram membrane protein. Penicillins and cephalosporins competed for the same binding sites. The antibiotic concentrations which excluded [14C]benzylpenicillin from 50% of the binding sites were those which inhibited by 50% the membrane-bound transpeptidase. The same rate constant (about 1 × 10−4 s−1) for the dissociation of the benzylpenicillin membrane complex at 37°C and in 0.017 M K2HPO4, was calculated either from the release of the radioactivity (using [14C]benzylpenicillin) or from the recovery of the transpeptidase activity. These observations supported the conclusion that the high-affinity binding sites in the isolated membranes were the transpeptidase molecules. All the complexes formed between the membranes and the various penicillins and cephalosporins examined were reversible at 37°C and in 0.017 M K2HPO4 at least with regard to the transpeptidase. Depending upon the antibiotics, the rate constants for the dissociation of these complexes varied from 3.3 × 10−3 to 0.73 × 10−4 s−1. The radioactivity released through the dissociation of [14C]benzylpenicillin membrane complex occurred mainly in the form of a compound which behaved as [W]-benzylpenicilloic acid both by paper electrophoresis and thin-layer chromatography. It was impossible to choose between several possible mechanisms for the release of the antibiotic molecule. [less ▲]

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See detailMembrane-bound DD-carboxypeptidase and LD-transpeptidase of Streptococcus faecalis ATCC 9790
Coyette, Jacques; Perkins, Harnold R.; Polacheck, Itzhack et al

in European Journal of Biochemistry (1974), 44(2), 459-468

Isolated membranes of Streptococcus faecalis ATCC 9790 exhibit DD-carboxypeptidase activity (standard reaction: Ac2-l-Lys-d-Ala-d-Ala →d-alanine + Ac2-l-Lys-d-Ala) and ld-trans-peptidase activity ... [more ▼]

Isolated membranes of Streptococcus faecalis ATCC 9790 exhibit DD-carboxypeptidase activity (standard reaction: Ac2-l-Lys-d-Ala-d-Ala →d-alanine + Ac2-l-Lys-d-Ala) and ld-trans-peptidase activity (standard reaction: Ac2-l-Lys-d-Ala + acceptor →d-alanine + Ac2-l-Lys-acceptor). The DD-carboxypeptidase activity has a considerable specificity for peptides with a C-terminal l-R3-d-Ala-d-Ala sequence where R3 is an amino acid residue and a long side-chain at the l-R3 position. A corresponding DD-transpeptidation reaction yielding the product Ac2-l-Lys-d-Ala-d-[14C]Ala from the system Ac2-l-Lys-d-Ala-d-Ala-f-d-[14C] alanine was not detected. The ld-transpeptidase activity has a considerable specificity for peptide donors that have an Nα-substituted, C-terminal l-R3-d-Ala sequence with a free ω-amino group at the end of a long side-chain at the l-R3 position, and a considerable specificity for amino group acceptors that are located on a d-carbon in α-position to a free carboxyl group. In the absence of acceptor, hydrolysis of the dipeptide Ac2-l-Lys-d-Ala (ld-carboxypeptidase activity) was not observed. Both DD-carboxypeptidase and ld-transpeptidase activities are inhibited by β-lactam antibiotics, but their relative sensitivity differs according to the particular antibiotic used. [less ▲]

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