ABSTRACT
Background & objectives : Acinetobacter baumannii is a Gram-negative, cocco-bacillus aerobic pathogen responsible for nosocomial infections in hospitals. In the recent past A. baumannii 0had developed resistance against β-lactams, even against carbapenems. Penicillin-binding proteins (PBPs) are crucial for the cell wall biosynthesis during cell proliferation and these are the target for β-lactams. Therefore, the present study was carried out to identify the PBPs in three (low, intermediate and high MICs) groups of carbapenem resistant isolates strains of A. baumannii. Methods: ATCC 19606 and 20 β-lactam resistant isolates of A. baumannii were obtained. Selective identification of the PBPs was done using Bocillin FL, a non-radioactive fluorescent derivative of penicillin. Results: The fluorescence emission from Bocillin-tag in SDS-PAGE gel of native strain identified eight PBPs, with apparent molecular weight of 94, 65, 49, 40, 30, 24, 22 and 17 kDa, however, these PBPs revealed alteration in carbapenem-resistant isolates. Interpretation & conclusions: A comparative analysis of PBPs in the resistant isolates with those of ATCC revealed a decreased expression of all PBPs except that of 65 and 17 kDa PBPs which were marginally downregulated, and simultaneous appearance of new 28 kDa PBP (in low and intermediate resistant isolates) and 36 kDa in high meropenem resistant group of A. baumannii. The present study indicated an association between alteration in PBPs and β-lactam resistance in A. baumannii.
Subject(s)
Acinetobacter baumannii/drug effects , Acinetobacter baumannii/metabolism , Amino Acid Sequence , Anti-Bacterial Agents/pharmacology , Carbapenems/pharmacology , Drug Resistance, Microbial , Electrophoresis, Polyacrylamide Gel , Microbial Sensitivity Tests , Molecular Sequence Data , Penicillin-Binding Proteins/chemistry , Penicillin-Binding Proteins/metabolism , Spectrometry, Mass, Matrix-Assisted Laser Desorption-IonizationABSTRACT
The quinolones exert their anti-bacterial activity by binding to DNA gyrase A (GyrA), an essential enzyme in maintenance of DNA topology within bacterial cell. The mutations conferring resistance to quinolones arise within the quinolone-resistance-determining region (QRDR) of GyrA. Therefore, quinolones interaction with wild and mutated GyrA can provide the molecular explanation for resistance. Resistant strains of Salmonella enterica of our hospital have shown mutations in the QRDR of GyrA of serine 83 (to phenylalanine or tyrosine) or aspartic acid 87 (to glycine or tyrosine). In order to understand the association between observed resistance and structural alterations of GyrA with respect to quinolone binding, we have studied the interaction of mutated QRDR of GyrA with nalidixic acid and ciprofloxacin by molecular modeling using GLIDE v4. Analysis of interaction parameters like G-score has revealed reduced interaction between nalidixic acid/ciprofloxacin with QRDR of GyrA in all four mutated cases of resistant strains. The mutation of Ser83 to Phe or Tyr shows least binding for nalidixic acid, while Asp87 to Gly or Tyr exhibits minimal binding for ciprofloxacin. The study also highlights the important role of arginines at 21, 91 and His at 45, which form strong hydrogen bonds (at < 3 Å) with quinolones. The hydrophilic OH group of Serine 83, which is in close proximity to the quinolone binding site is replaced by aromatic moieties of Tyr or Phe in mutated GyrA. This replacement leads to steric hindrance for quinolone binding. Therefore, quinolone resistance developed by Salmonella appears to be due to the decreased selectivity and affinity of nalidixic acid/ciprofloxacin to QRDR of GyrA.