Phenylalkylsulfonyl derivatives as covalent inhibitors of penicillin amidase

It was demonstrated that phenylmethanesulfonyl fluoride-a very potent inhibitor of penicillin amidase from Escherichia coli-binds covalently to the enzyme in molar ratio 1:1. The chloride, the azide and the N-hydroxysuccinimide ester of phenylmethanesulfonic acid are also very strong inactivators of the amidase. Weaker inhibition was noted with para-substituted phenylmethanesulfonyl chlorides and with phenylethanesulfonyl and alkylsulfonyl chlorides. The inactivated amidase could be reactivated by incubation either with 6-amino-penicillanic acid or with proteins from E. coli extract. Benzyl isocyanate is also a potent covalent inhibitor of the amidase but inactivated amidase could be not reactivated in this way. It was demonstrated that representatives of all inactivator types bind to one active site of the amidase. Interdependence between inactivation rate and stability of some sulfonyl inhibitors was observed. No inhibition was noted the amide, the hydrazide and the methyl ester of phenylmethanesulfonic acid.     

Synergistic action of combinations of penicillin and perhydroacridine derivatives

The effect of combinations of perhydroacridine derivatives such as 10-nitroso-trans-anti-cys-perhydroacridine (MT-2) and 10-isopropylamino-trans-anti-cys-perhydroacridine (MT-6) with benzylpenicillin on growth of two penicillinase-producing strains of Staphylococcus aureus was studied. The combination components were added to the culture simultaneously or at different periods i.e. the second component was added after preliminary exposure of the bacterial cells to the first component for 1-6 hours at 37 degrees C. It was shown that duration and efficacy of the combination synergistic action were directly proportional to the time of the component addition. The highest synergistic action of the combinations was observed when both the components were simultaneously added to the staphylococcal culture. The combinations were less efficient when the bacterial cells were preliminarily incubated with the perhydroacridines. Addition of the perhydroacridine derivatives after the strain contact with the penicillin resulted in elimination of the combination synergistic action. Thin-layer chromatography did not reveal complexing between the penicillin and perhydroacridines.     

Experimental and simulation studies reveal mechanism of action of human defensin derivatives

Antimicrobial peptides (AMPs) are naturally occurring promising candidates which can be used as antibiotics against a wide variety of bacteria. The key component for using them as a potent antibiotic is that their mechanism of action is less prone to bacterial resistance. However, the molecular details of their mechanism of action is not yet fully understood. In this study, we try to shed light on the mode of action of AMPs, possible reason behind it, and their interaction with lipid bilayers through experimental as well as molecular dynamics (MD) simulation studies. The focal of our study was Human beta defensin 3 (hBD-3) which is a naturally occurring AMP. We chose three derivatives of hBD-3, namely CHRG01, KSR, and KLR for the detailed analysis presented in this study. These three peptides are evaluated for their antibacterial potency, secondary structure analysis and mechanism of action. The experimental results reveal that these peptides are active against gram positive as well as gram negative bacteria and kill bacteria by forming membrane pores. The MD simulation results correlate well with the antibacterial activity and shed light into the early membrane insertion dynamics. Moreover, the specific amino acids responsible for membrane disruptions are also identified from the MD simulations. Understanding the molecular level interaction of individual amino acids with the lipid bilayer will greatly help in the design of more efficient antimicrobial peptides.     

QSAR studies on acylated histamine derivatives

H(3)-receptor antagonists activity in terms of -log K(i) for a series of acylated histamine derivatives was modeled using topological indices, namely negentropy (N), molecular redundancy (MRI), and valence connectivity index ((m)x(v)) indices. Excellent results were obtained in multiple regression analysis upon the introduction of a dummy parameter (indicator parameter). Consistent increase in R(2)(A) value indicated that inspite of observed collinearity the proposed models are significant.     

Molecular modeling studies of substrate binding by penicillin acylase

Molecular modeling has revealed intimate details of the mechanism of binding of natural substrate, penicillin G (PG), in the penicillin acylase active center and solved questions raised by analysis of available X-ray structures, mimicking Michaelis complex, which substantially differ in the binding pattern of the PG leaving group. Three MD trajectories were launched, starting from PDB complexes of the inactive mutant enzyme with PG (1FXV) and native penicillin acylase with sluggishly hydrolyzed substrate analog penicillin G sulfoxide (1GM9), or from the complex obtained by PG docking. All trajectories converged to a similar PG binding mode, which represented the near-to-attack conformation, consistent with chemical criteria of how reactive Michaelis complex should look. Simulated dynamic structure of the enzyme-substrate complex differed significantly from 1FXV, resembling rather 1GM9; however, additional contacts with residues bG385, bS386, and bN388 have been found, which were missing in X-ray structures. Combination of molecular docking and molecular dynamics also clarified the nature of extremely effective phenol binding in the hydrophobic pocket of penicillin acylase, which lacked proper explanation from crystallographic experiments. Alternative binding modes of phenol were probed, and corresponding trajectories converged to a single binding pattern characterized by a hydrogen bond between the phenol hydroxyl and the main chain oxygen of bS67, which was not evident from the crystal structure. Observation of the trajectory, in which phenol moved from its steady bound to pre-dissociation state, mapped the consequence of molecular events governing the conformational transitions in a coil region a143-a146 coupled to substrate binding and release of the reaction products. The current investigation provided information on dynamics of the conformational transitions accompanying substrate binding and significance of poorly structured and flexible regions in maintaining catalytic framework.