Role of β-lactam carboxyl group on binding of penicillins and cephalosporins to class C β-lactamases (original) (raw)
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Structural Aspects for Evolution of β-Lactamases from Penicillin-Binding Proteins
Journal of the American Chemical Society, 2003
Penicillin-binding proteins (PBPs), biosynthetic enzymes of bacterial cell wall assembly, and -lactamases, resistance enzymes to -lactam antibiotics, are related to each other from an evolutionary point of view. Massova and Mobashery (Antimicrob. Agents Chemother. 1998, 42, 1-17) have proposed that for -lactamases to have become effective at their function as antibiotic resistance enzymes, they would have had to undergo structure alterations such that they would not interact with the peptidoglycan, which is the substrate for PBPs. A cephalosporin analogue, 7 -[N-Acetyl-L-alanyl-γ-D-glutamyl-L-lysine]-3-acetoxymethyl-3-cephem-carboxylic acid (compound 6), was conceived and synthesized to test this notion. The X-ray structure of the complex of this cephalosporin bound to the active site of the deacylation-deficient Q120L/Y150E variant of the class C AmpC -lactamase from Escherichia coli was solved at 1.71 Å resolution. This complex revealed that the surface for interaction with the strand of peptidoglycan that acylates the active site, which is present in PBPs, is absent in the -lactamase active site. Furthermore, insertion of a peptide in the -lactamase active site at a location where the second strand of peptidoglycan in some PBPs binds has effectively abolished the possibility for such interaction with the -lactamase. A 2.6 ns dynamics simulation was carried out for the complex, which revealed that the peptidoglycan surrogate (i.e., the active-site-bound ligand) undergoes substantial motion and is not stabilized for binding within the active site. These factors taken together disclose the set of structure modifications in the antibiotic resistance enzyme that prevent it from interacting with the peptidoglycan, en route to achieving catalytic proficiency for their intended function.
Computational analysis of the interactions of a novel cephalosporin derivative with β-lactamases
BMC Structural Biology
Background: One of the main concerns of the modern medicine is the frightening spread of antimicrobial resistance caused mainly by the misuse of antibiotics. The researchers worldwide are actively involved in the search for new classes of antibiotics, and for the modification of known molecules in order to face this threatening problem. We have applied a computational approach to predict the interactions between a new cephalosporin derivative containing an additional β-lactam ring with different substituents, and several serine β-lactamases representative of the different classes of this family of enzymes. Results: The results of the simulations, performed by using a covalent docking approach, has shown that this compound, although able to bind the selected β-lactamases, has a different predicted binding score for the two βlactam rings, suggesting that one of them could be more resistant to the attack of these enzymes and stay available to perform its bactericidal activity. Conclusions: The detailed analysis of the complexes obtained by these simulations suggests possible hints to modulate the affinity of this compound towards these enzymes, in order to develop new derivatives with improved features to escape to degradation.
Biochemistry, 2010
Penicillin-binding proteins (PBPs) are the molecular target for the widely used β-lactam class of antibiotics, but how these compounds act at the molecular level is not fully understood. We have determined crystal structures of E. coli PBP5 as covalent complexes with imipenem, cloxacillin and cefoxitin. These antibiotics exhibit very different second order rates of acylation for the enzyme. In all three structures, there is excellent electron density for the central portion of the βlactam, but weak or absent density for the R1 or R2 side chains. Areas of contact between the antibiotics and PBP 5 do not correlate with the rates of acylation. The same is true for conformational changes because although shift of a loop leading to an electrostatic interaction between Arg248 and the β-lactam carboxylate, which occurs completely with cefoxitin, partially with imipenem and is absent with cloxacillin, is consistent with the different rates of acylation, mutagenesis of Arg248 only decreased cefoxitin acylation two fold. Together, these data suggest that structures of post-covalent complexes of PBP 5 are unlikely to be useful vehicles for design of new covalent inhibitors of PBPs. Finally, superimposition of the imipenem-acylated complex with PBP5 in complex with a boronic acid peptidemimetic shows that the position corresponding to the hydrolytic water molecule is occluded by the ring nitrogen of the β-lactam. Since the ring nitrogen occupies a similar position in all three complexes, this supports the hypothesis that deacylation is blocked by the continued presence of the leaving group after opening of the β-lactam ring. Penicillin-binding proteins (PBPs) are so named because they are the targets for the wellknown and widely used class of β-lactam antibiotics. Many PBPs are transpeptidases (TPases) that catalyze the formation of cross-links between peptides on adjacent strands of glycan during the final stages of peptidoglycan synthesis in bacteria, thereby conferring strength to the cell wall against osmotic pressure. Other PBPs are carboxypeptidases (CPases), which remove the terminal D-Ala of the pentapeptidyl substrate, or endopeptidases, † This work was supported by the National Institutes of Health grants GM66861 to CD, AI36901 to RAN and AI17986 to RFP.
Proceedings of the National Academy of Sciences, 1993
The structure of the class C ampC beta-lactamase (cephalosporinase) from Enterobacter cloacae strain P99 has been established by x-ray crystallography to 2-A resolution and compared to a class A beta-lactamase (penicillinase) structure. The binding site for beta-lactam (penicillinase) structure. The binding site for beta-lactam antibiotics is generally more open than that in penicillinases, in agreement with the ability of the class C beta-lactamases to better bind third-generation cephalosporins. Four corresponding catalytic residues (Ser-64/70, Lys-67/73, Lys-315/234, and Tyr-150/Ser-130 in class C/A) lie in equivalent positions within 0.4 A. Significant differences in positions and accessibilities of Arg-349/244 may explain the inability of clavulanate-type inhibitors to effectively inactivate the class C beta-lactamases. Glu-166, required for deacylation of the beta-lactamoyl intermediate in class A penicillinases, has no counterpart in this cephalosporinase; the nearest candidate, Asp-217, is 10 A from the reactive Ser-64. A comparison of overall tertiary folding shows that the cephalosporinase, more than the penicillinase, is broadly similar to the ancestral beta-lactam-inhibited enzymes of bacterial cell wall synthesis. On this basis, it is proposed that the cephalosporinase is the older of the two beta-lactamases, and, therefore, that a local refolding in the active site, rather than a simple point mutation, was required for the primordial class C beta-lactamase to evolve to the class A beta-lactamase having an improved ability to catalyze the deacylation step of beta-lactam hydrolysis.
Journal of Pharmacy and Pharmacology 4 (2016) 212-225, 2016
An approach of using molinspiration calculations and molecular docking on PBPs (penicillin-binding proteins) and certain β-lactamases is employed to predict the molecular properties, bioactivity and resistance of newer and reference cephalosporins. The previously synthesized cephalosporins 1-8 and reference cephalosporins were subjected to extensive evaluations by calculating the molecular properties, drug-likeness scores on the bases of Lipinski's rule and bioactivity prediction using the method of molinspiration web-based software. The TPSA (topological polar surface area), OH-NH interactions, n-violation and the molinspiration Log partition coefficient (miLogP) values were also calculated. The investigated cephalosporins were subjected to molecular docking study on PBPs (1pyy) and on β-lactamases produced by S. aureus, K. pneumonia, E. coli and P. auroginosa using 1-click-docking website. Molecular properties of 1-8 recorded higher TPSA than cephalexin and were lower than the reference cephalosporins and do not fulfill the requirements for Lipinski's rule. Bioactivities of 1-8 were predicted to be less and their docking scores on PBPs were comparable to those of the reference cephalosporins, particularly ceftobiprole. The references recorded various docking scores on the above β-lactamases and as expected, ceftobiprole recorded the lowest scores on all β-lactamases. Cephalosporins 1-8 recorded various docking scores on β-lactamases. Molecular docking studies on PBPs and β-lactamases are considered as very useful, reliable and practical approach for predicting the bioactivity scores and to afford some information about the stability and selectivity of the newly proposed cephalosporins against β-lactamases of certain pathogenic microbes, such as P. auroginosa and MRSA, by recording the relative docking scores in comparison with those of reference cephalosporins.
Molecular Dynamics Simulations of the TEM-1 β-Lactamase Complexed with Cephalothin
Journal of Medicinal Chemistry, 2005
Herein, we present theoretical results aimed at elucidating the origin of the kinetic preference for penicillins over cephalosporins characteristic of the TEM/SHV subgroup of class A -lactamases. First, we study the conformational properties of cephalothin showing that the C2-down conformer of the dihydrothiazine ring is preferred over the C2-up one by ∼2 kcal/mol in solution (0.4-1.4 kcal/mol in the gas phase). Second, the TEM-1 -lactamase complexed with cephalothin is investigated by carrying out a molecular dynamics simulation. The ∆G binding energy is then estimated using molecular mechanics Poisson-Boltzmann surface area (MM-PBSA) and quantum chemical PBSA (QM-PBSA) computational schemes. The preferential binding of benzylpenicillin over cephalothin is reproduced by the different energetic calculations, which predict relative ∆∆G binding energies ranging from 1.8 to 5.7 kcal/mol. The benzylpenicillin/ cephalothin ∆∆G binding energy is most likely due to the lower efficacy of cephalosporins than that of penicillins in order to simultaneously bind the "carboxylate pocket" and the "oxyanion hole" in the TEM-1 active site.
Biochemistry, 1995
Two clinically-important beta-lactam antibiotics, cephalothin and cefotaxime, have been observed by X-ray crystallography bound to the reactive Ser62 of the D-alanyl-D-alanine carboxypeptidase/transpeptidase of Streptomyces sp. R61. Refinement of the two crystal structures produced R factors for 3 sigma (F) data of 0.166 (to 1.8 A) and 0.170 (to 2.0 A) for the cephalothin and cefotaxime complexes, respectively. In each complex, a water molecule is within 3.1 and 3.6 A of the acylated beta-lactam carbonyl carbon atom, but is poorly activated by active site residues for nucleophilic attack and deacylation. This apparent lack of good stereochemistry for facile hydrolysis is in accord with the long half-lives of cephalosporin intermediates in solution (20-40 h) and the efficacy of these beta-lactams as inhibitors of bacterial cell wall synthesis. Different hydrogen binding patterns of the two cephalosporins to Thr301 are consistent with the low cefotaxime affinity of an altered penicillin-binding protein, PBP-2x, reported in cefotaxime-resistant strains of Streptococcus pneumoniae, and with the ability of mutant class A beta-lactamases to hydrolyze third-generation cephalosporins.
Chemistry & Biodiversity, 2005
We report a molecular-mechanics (AMBER*) study on the Henry ± Michaelis complex and the corresponding acyl ± enzyme adduct formed between imipenem (1), a transient inhibitor of b-lactamases, and Enterobacter cloacae P99, a class C-b-lactamase. We have examined the influence of the structural configuration of the functional groups in the substrate on their three-dimensional (3D) arrangement at the active site, which was compared with those adopted by typical penicillins and cephalosporins. Our results confirm that the carboxy group of the antibiotic plays a prominent role in the binding of the substrate to the active site, and that it activates Ser 64 through interaction with the phenolic OH group of Tyr 150 . The binding of imipenem to E. cloacae P99 increases the distance between Tyr 150 and Ser 64 due to the presence of a hydrophobic Me group in the (R)-1hydroxyethyl substituent at C(6). This, together with the 3D arrangement of its carboxy group, leads to an interaction with the active site in a manner that hinders H exchange between the nucleophile in Ser 64 and its basic activator, the phenolic group of Tyr 150 .