Conformation–activity relationships and the mechanism of action of penicillin

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The docking of penicillins to a computer model of the active site of the penicillin-binding protein of Streptornyces R61 produces structures that exhibit a four-centred interaction between the 0 -H of a serine residue and the (0)C-N of the p-lactam ring. If such a structure is a stationary point on the reaction coordinate for the acylation of the serine OH by the p-lactam carbonyl group, the acylation mechanism would appear to consist of 0-C and H-N bond formation, concerted with the cleavage of the C-N bond. The existence of this "N-protonation" mechanism, and the energetics of this mechanism in comparison to the usual "0-protonation" pathway in which proton transfer to oxygen and addition to the carbonyl group precede C-N bond fission, have been examined by ab initio M O calculations on the neutral hydrolysis and methanolysis of N-methylazetidinone, penam, and penam-3a-carboxylate. The geometries of reactants and transition structures have been optimized fully at the 3-21G or 3-21G* levels, the existence of transition structures has been confirmed by vibrational analysis, thermochemical data have been computed, and the one-point energies of all structures have been determined at the MP216-31G* level. The N-protonation mechanism is found to exist with all substrates, and to be preferred over the 0-protonation mechanism by over 5 kcallmol. The activation energies are 5 kcallmol lower in the bicyclic penam than in the monocyclic N-methylazetidinone, and attack from the convex face is preferred over attack from the concave face. The introduction of a 3a-carboxylate group, as in penicillin itself, results in an additional 5 kcallmol decrease in activation energy. The origins of these trends are discussed. Since the active sites of some penicillin-recognizing enzymes contain at least one water molecule, catalysis of the foregoing reactions by one water molecule has also been examined. The catalysis amounts to over 10 kcaVmol in both the N-and 0-protonation mechanisms, and the preference for the N-protonation mechanism is maintained. It is concluded that penicillin complexes to its receptor in such a manner as to allow acylation of the active site serine residue to proceed via the energetically most favourable mechanism. SAUL WOLFE, CHAN-KYUNG KIM et KIYULL YANG. Can. J. Chem. 72, 1033Chem. 72, (1994. L'ammage de pCnicillines au modble sur ordinateur du site actif de la protCine de liaison de la pCnicilline du Streptomyces R61 foumit des structures qui prCsentent une interaction 21 quatre centres entre le 0 -H de la sCrine et le (0)C-N du noyau p-lactame. Si une telle structure est un point stationnaire sur les coordonnCes rtactionnelles de l'acylation du OH de la sCrine par le groupe carbonyle du p-lactame, il semblerait que le mtcanisme d'acylation cornporterait la formation de liaisons 0-C et H-N concertCe avec un clivage de la liaison C-N. L'existence de ce mCcanisme de ccN-protonation, et les tnergies impliquCes dans ce mCcanisme, par comparaison avec la voie usuelle de ccO-protonation,, dans laquelle le trans...

Section: Catalysis By Water Of the Hydrolysis And Methanolysis Of 3 And5mentioning

confidence: 59%

Ab initio molecular orbital calculations on the neutral hydrolysis and methanolysis of azetidinones, including catalysis by water. Relationship to the mechanism of action of β-lactam antibiotics

Wolfe¹,

Kim²,

Yang³

1994

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“…We have recently discussed the abilities of penicillins (1) and cephalosporins (2) to "fit" to a com uter-generated model ( (4). This peptide contains the residues that surround the active site serine of a PBP from Streptomyces R6 1, which has been studied in some detail (5).…”

Section: Theoretical Considerations: "Fit" and "Reactivity"mentioning

confidence: 99%

Interactive design and synthesis of a novel antibacterial agent

Wolfe¹,

Jin²,

Yang³

et al. 1994

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. Can. J. Chem. 72, 105 1 (1994). P-Lactam compounds act on penicillin-recognizing enzymes via acylation of the hydroxyl group of an active site serine. When the resulting acyl enzyme is kinetically stable, as in the case of a penicillin-binding protein (PBP), the biosynthesis of a bacterial cell wall is inhibited, and death of the organism results. The de novo design of an antibacterial agent targeted to a PBP might be possible if the three-dimensional structural requirements of the equilibrium (i.e.,fit) and catalytic (i.e. reactivity) steps of the aforementioned enzymatic process could be determined. For a model of the active site of a PBP from Streptomyces R61, the use of molecular mechanics calculations to treat "fit," and ab initio molecular orbital calculations to treat "reactivity," leads to the idea that the carboxyl group (GI) and the amide N-H (G2) of the antibiotic are hydrogen bonded to a lysine amino group and a valine carbonyl group in the enzyme-substrate complex. These two hydrogen bonds place the serine hydroxyl group on the convex face of the antibiotic, in position for attack on the p-lactam ring by a neutral reaction, catalyzed by water, that involves a direct proton transfer to the p-lactam nitrogen. Molecular orbital calculations of structure-reactivity relations associated with this mechanism suggest that C=N is bioisosteric to the p-lactam N-C(=O), comparable to a p-lactam in its reactivity with an alcohol, and that the product RO(C-N)H is formed essentially irreversibly (-AE > 10 kcaUmo1). Accordingly, structures containing a G1 and a G2 separated by a C=N, and positioned in different ways with respect to this functional group, have been synthesized computationally and examined for their ability to fit to the PBP model. This strategy identified a 2H-5,6-dihydro-l,4-thiazine substituted by hydroxyl and carboxyl groups as a target for chemical synthesis. However, exploratory experiments suggested that the C=N of this compound equilibrates with endocyclic and exocyclic enamine tautomers. This required that the C2 position be substituted, and that the hydroxyl group not be attached to the carbon atom adjacent to the C=N. These conditions are met in a 2,2-dimethyl-3-(2-hydroxypropy1)-1,4-thiazine, which also exhibits the necessary fit to the PBP model. Two epimers of this compound have been synthesized, from D-and L-serine. The compound derived from L-serine is not active. The compound derived from D-serine exhibits antibacterial activity, but is unstable, and binding studies with PBP's have not been performed. It is hoped that these studies can be canied out if modification of the lead structure leads to compounds with improved chemical stability. Toutefois, des essais prkliminaires ont suggCrC que le C=N de ce composC serait en Cquilibre avec les Cnamines endocycliques et exocycliques tautombres. Cette caractkristique nCcessite que la position C2 soit substituCe et que le groupe hydroxyle ne soit pas attach6 au carbone adjacent du C=N. On rencontre ces conditions dans une 2,2-dimCthy1-3-(2...

“…Figure 2 illustrates this point. Structure A (4) is a closeup view of the C-0-WH-0-HI N-C=O geometry in the complex formed between the crystal structure of penicillin G (11), a water molecule, and 12, a hexapeptide model of the penicillin receptor (3). To form this complex, G I , the carboxyl group of the antibiotic, was hydrogen bonded to the terminal amino group of a lysine residue of 12, labelled N, and G2, the side-chain N-H of the antibiotic, was hydrogen bonded to a carbonyl oxygen of 12, labelled V Structure B (4) is a closeup view of the C-0-WH-0-WN-C=O geometry in the ab initio (3-21G) complex that proceeds to the transition state of the water-catalyzed methanolysis of N-methylazetidinone.…”

Section: Theoretical Considerationsmentioning

confidence: 99%

“…of 1. Sixteen initial orientations of GI (C02-) and G2 (O-H) of this structure with respect to the amino (N) and carbonyl (V) groups of 12 were minimized (3,4) to produce, as the lowest energy structure, the complex shown in Fig Ten conformations of the RWSS diastereomer and seven conformations of the RS/SR diastereomer of 1, corresponding to torsion about the C5-C6, C3-C7, C7-C8, and C8-0 bonds, were generated using the dihedral driver of M M P~ (85) (14), and the energies of these structures were minimized. Each of the 34 RR, SS, RS, and SR structures was then docked to 12, as described above, the energies of the docked structures were minimized, and the rms deviations from the four-centred relationship of Fig.…”

Section: Theoretical Considerationsmentioning

confidence: 99%

Synthesis of the stereoisomers of a novel antibacterial agent and interpretation of their relative activities in terms of a theoretical model of the penicillin receptor

Wolfe¹,

Zhang²,

Johnston³

et al. 1994

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.2,2-~imeth~l-3-(2'-h~drox~~ro~~l)-5-carbox~-~~-1,4-thiazine (1) is a designed antibacterial agent. Based on an analysis of how penicillin complexes to and reacts with a model of a penicillin-binding protein, 1 contains a functional group (C=N) that can react with a serine hydroxyl group of the receptor according to the putative reaction Enz-OH + C=N + Enz-0-C-NH. Compound 1 also contains additional substituents that are designed to position the 0-H and C=N groups relative to one another in the enzyme-substrate complex in a geometry that attempts to reproduce the optimum geometry of approach of two such reactants. A most important assumption is that this optimum geometry can be computed ab initio. In a first preparation of 1, (?)-5-methyl-4-hexene-2-ol(2) was converted to the lithium salt of (?)-2-mercapto-2-methy1-5-tert-butyldimethylsiloxy-3-hexanone (7), which was condensed with the N-tert-butoxycarbonyl-D-and L-serine-p-lactones (3). The synthesis was completed by deprotection with formic acid and cyclization in water. The R and S enantiomers of 2 have now been obtained, and the absolute configuration of the alcohol established, by reaction of the R-and S-propylene oxides with an organometallic reagent prepared from p,p-dimethylvinyl bromide. The R alcohol has also been secured by lipase-catalyzed transesterification with trifluoroethyl butyrate, and chemical hydrolysis of the trifluoroethyl ester. The R and S enantiomers of 2 were converted to the R and S enantiomers of 7, and these were condensed with the R and S enantiomers of 3 to yield each of the stereoisomers of the chemically unstable 1 in ca. 95% optically pure form. Antibacterial activity resides in the 5S,8R and 5S,8S isomers. These findings are shown to be consistent with the theoretical model. It is hoped that the stability of the lead structure 1 can be improved, to allow binding experiments with penicillin recognizing enzymes to proceed.SAUL WOLFE, C m ZHANG, BLAIR D. JOHNSTON et CHAN-KYUNG KIM. Can. J. Chem. 72, 1066Chem. 72, (1994.La 2,2-dimCthyl-3-(2'-hydroxypropyl)-5-c~3-1,4-thiazine (1) devrait &tre une agent antibactkrien. Sur la base d'une analyse du mCcanisme de complexation et de reaction de la ptnicilline avec un modkle d'une protkine se liant B la pknicilline, 1 contient un groupe fonctionnel (C=N) qui peut rCagir avec un groupe hydroxyle de la strine du recepteur suivant la reaction Enz-OH + C=N + Enz-0-C-NH. Le composC 1 contient aussi des substituants additionnels choisis pour orienter les groupes 0 -H et C=N I'un par rapport B l'autre dans le complexe enzyme-substrat dans une g6omCtrie qui essaye de reproduire la gComCtrie optimale de I'approche de deux tels riactants. Une hypothkse trks importante est que cette gCom6trie optimale peut &tre calculCe ab initio. Dans une premikre preparation du composC 1, on a transform6 le (?)-5-methyl-hex-4-kn-2-ol(2) en sel de lithium de la (~)-2-mercapto-2-mCthyI-5-tert-butyldimCthylsiloxyhexan-3-one (7) qui a Ct C condensee avec les N-tert-butoxycarbonyl-D-et L-s6rine-p-lactones (3). On a c...