Penicillin-Resistant Mechanisms in Pseudomonas aeruginosa : Binding of Penicillin to Pseudomonas aeruginosa KM 338

Abstract

A comparison of the binding of radioactive penicillin G to whole cells and the membrane fraction derived from Pseudomonas aeruginosa KM 338 was made. This organism has intrinsic resistance to penicillin. The binding to the membrane fraction which catalyzed peptidoglycan synthesis followed saturation type kinetics and saturation was achieved at approximately 2 nmol of penicillin G per ml, whereas binding to the whole cells was entirely of the nonsaturation type. The binding of carbenicillin to the membrane fraction was determined by competition between radioactive penicillin G and unlabeled carbenicillin for the binding sites. It was bound at the same sites in almost the same manner. When whole cells were pretreated with high concentration of unlabeled penicillin G or carbenicillin, the subsequent binding of radioactive penicillin G to the membrane fraction from carbenicillin-treated cells was entirely nonspecific, but with penicillin G-pretreated cells it was still specific. There was apparently specific binding of radioactive penicillin G to ethylenediaminetetraacetate-treated cells. P. aeruginosa KM 338 had an extremely low activity of β-lactamase compared with other enzyme-producing organisms. This enzyme from P. aeruginosa KM 338 was of the cephalosporinase type. These data indicate that penicillin resistance of P. aeruginosa KM 338 may be a consequence of the development of a permeability barrier which prevents the antibiotic from reaching its sites of action in the cytoplasmic membrane.

Prevention of penicillin-induced lysis of Staphylococcus aureus by cellular lipoteichoic acid.

“…The lipoteichoic acid inhibited either extracellular autolysin activity in culture supernatant or autolysis of whole cells in exponential phase of S. aureus. These results indicate that the prevention of penicillin-induced lysis of S. aureus by the lipoteichoic acid was brought about by the inhibition of autolytic activity of the organism.It has been well known that /3-lactam antibiotics are specifically and irreversibly bound to their target sites on the cytoplasmic membrane of Gram-positive1~4) and 3,5) where they inhibit cross-linking reaction in peptidoglycan synthesis6~11)The target sites are the penicillin binding proteins, transpeptidase and D-alanine carboxypeptidase12). Until recently, the bacteriolysis has been assumed to be induced by the binding of 8-lactam antibiotics to the penicillin binding proteins, indicating that the lysis was a consequence of the inhibition of cell wall biosynthesis2,13).…”

“…It has been well known that /3-lactam antibiotics are specifically and irreversibly bound to their target sites on the cytoplasmic membrane of Gram-positive1~4) and Gram-negative bacteria2, 3,5) where they inhibit cross-linking reaction in peptidoglycan synthesis6~11)The target sites are the penicillin binding proteins, transpeptidase and D-alanine carboxypeptidase12). Until recently, the bacteriolysis has been assumed to be induced by the binding of 8-lactam antibiotics to the penicillin binding proteins, indicating that the lysis was a consequence of the inhibition of cell wall biosynthesis2,13).…”

Effect of Piperacillin on D-Alanine Carboxypeptidase Activities from Pseudomonas aeruginosa

“…The organism used was Pseudomonas aeruginosa KM338, derived from ATCC 17653 ; this was the same strain as used in our previous study (Suginaka et al, 1975) of penicillin-resistant mechanisms. As estimated by a serial tube dilution method in Trypticase Soy Broth (BBL) with an inoculum of about lo4 bacteria ml-l, the minimal inhibitory concentrations of piperacillin, carbenicillin and benzylpenicillin were 12.5, 125 and 30000 pg ml-l, respectively.…”

Comparison of the Binding of Penicillin G to Staphylococcal L-Form and Its Parent Strain Membranes

“…It seems likely that this organism must lack some enzyme(s) of the cell wall biosynthetic pathway leading to cross-linking of peptidoglycan. It has been suggested that penicillin is specifically and irreversibly bound to transpeptidase in the cytoplasmic membrane of penicillin-susceptible (1,7,8) and relatively unsusceptible organisms, such as Pseudomonas.aeruginosa (10), and consequently inhibits the cross-linking reaction (1,7,9). The purpose behind this report was to determine the penicillin-binding ability of the isolated membrane from a strain of staphylococcal L-form, compared with that from its parent strain.…”

“…The membrane fraction was obtained from the washed cell pellet by osmotic lysis, sonic treatment, and deoxyribonuclease digestion. The parent strain was grown in brain heart infusion broth (BBL) for 4.5 h (midlog phase), and the membrane fraction was prepared from the washed cells after lysis with cell wall lytic enzyme, ALE (11,12) Amersham/Searle, Buckinghamshire, England) as described previously (10).…”