Mixed Quantum Mechanical/Molecular Mechanical (QM/MM) Study of the Deacylation Reaction in a Penicillin Binding Protein (PBP) versus in a Class C β-Lactamase

Gherman, Benjamin F.; Goldberg, Shalom D.; Cornish, Virginia W.; Friesner, Richard A.

doi:10.1021/ja036879a

Cited by 76 publications

(122 citation statements)

References 85 publications

(170 reference statements)

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“…In a second model, the tyrosine acts as conjugate general base together with Lys67 (Figure 4a). 25 This hypothesis is more consistent with this structure and previous studies, though we do not fully favor it either. Rather, together with previous biochemical studies, our ultrahigh-resolution structure favors a third model where a phenol form of Tyr150 stabilizes the tetrahedral deacylation transition state in conjunction with the lactam nitrogen of the substrate (Figure 4b).…”

Section: Discussionsupporting

confidence: 79%

“…Lys315 is believed to be positively charged and it is likely that it functions as the donor in all three hydrogen bonds as well. 25 At a dihedral angle of 46.3°, the hydrogen bond between Lys315 and Tyr150 is more in plane with the phenol ring, compared with Lys67 whose Nζ is out of the plane by 82.2°. Thus, Tyr150 seems to form two hydrogen bonds in the active site, donating a hydrogen to the boronic acid O2 atom, which represents the position of the attacking deacylating water, and accepting a hydrogen from the Lys315 Nζ atom.…”

Section: Lys67mentioning

confidence: 99%

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The Deacylation Mechanism of AmpC β-Lactamase at Ultrahigh Resolution

et al. 2006

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Abstractβ-Lactamases confer bacterial resistance to β-lactam antibiotics, such as penicillins. The characteristic class C β-lactamase AmpC catalyzes the reaction with several key residues including Ser64, Tyr150, and Lys67. Here, we describe a 1.07 Å X-ray crystallographic structure of AmpC β-lactamase in complex with a boronic acid deacylation transition-state analogue. The high quality of the electron density map allows the determination of many proton positions. The proton on the Tyr150 hydroxyl group is clearly visible and is donated to the boronic oxygen mimicking the deacylation water. Meanwhile, Lys67 hydrogen bonds with Ser64Oγ, Asn152Oδ1, and the backbone oxygen of Ala220. This suggests that this residue is positively charged and has relinquished the hydrogen bond with Tyr150 observed in acyl-enzyme complex structures. Together with previous biochemical and NMR studies, these observations indicate that Tyr150 is protonated throughout the reaction coordinate, disfavoring mechanisms that involve a stable tyrosinate as the general base for deacylation. Rather, the hydroxyl of Tyr150 appears to be well positioned to electrostatically stabilize the negative charge buildup in the tetrahedral high-energy intermediate. This structure, in itself, appears consistent with a mechanism involving either Tyr150 acting as a transient catalytic base in conjunction with a neutral Lys67 or the lactam nitrogen as the general base. Whereas mutagenesis studies suggest that Lys67 may be replaced by an arginine, disfavoring the conjugate base mechanism, distinguishing between these two hypotheses may ultimately depend on direct determination of the pK a of Lys67 along the reaction coordinate.

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Section: Discussionsupporting

confidence: 79%

Section: Lys67mentioning

confidence: 99%

The Deacylation Mechanism of AmpC β-Lactamase at Ultrahigh Resolution

et al. 2006

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“…Moreover, for the majority of enzymes studied, structural differences between reactant and transition state are localized near catalytic residues, whose identities were often fixed. Ultimately, it is likely that attention to properties of electronic structure, for example, via mixed quantum͞molecular mechanics methods (24), will be required for full generality without heuristic assumptions regarding catalytic residues.…”

Section: Discussionmentioning

confidence: 99%

Sequence optimization and designability of enzyme active sites

Chakrabarti

Klibanov

Friesner

2005

Proc. Natl. Acad. Sci. U.S.A.

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We recently found that many residues in enzyme active sites can be computationally predicted by the optimization of scoring functions based on substrate binding affinity, subject to constraints on the geometry of catalytic residues and protein stability. Here, we explore the generality of this surprising observation. First, the impact of hydrogen-bonding networks necessary for catalysis on the accuracy of sequence optimization is assessed; incorporation of these networks, where relevant, into the set of catalytic constraints is found to be essential. Next, the impact of multiple substrate selectivity on sequence optimization is probed by carrying out independent calculations for complexes of deoxyribonucleoside kinases with various cognate ligands, revealing how simultaneous selection pressures determined active-site sequences of these enzymes. Including previous calculations on simpler enzymes, computational sequence optimization correctly predicts 76% of all active-site residues tested (86% correct, with 93% similar, for naturally conserved residues). In these studies, the ligand is fixed in its native conformation. To assess the applicability of these methods to de novo active-site design, the effect of small ligand motions around the native pose is also examined. Robustness of sequence accuracy for topologically similar poses is demonstrated for selected kinases, but not for a model peptidase. Based on these observations, we introduce the notion of the designability of an enzyme active site, a metric that may be used to guide the search for protein scaffolds suitable for the introduction of de novo activity for a desired chemical reaction.S ince it was first observed that all known natural proteins adopt one of only Ϸ1,000 distinct protein folds (1), researchers working in the field of computational protein design have endeavored to understand the basis of this biophysical phenomenon and apply the resultant knowledge to the construction of unnatural proteins. Motivated by the observation that certain folds are associated with more natural sequences than others, the principle of fold designability (the number of unique sequences compatible with the backbone structure of the fold) was introduced (1). To place the problem on a mathematical foundation, an analogy has been drawn between sequence space and phase space in statistical physics, where the backbone structure represents a macroscopic state of the system and distinct sequences are microscopic states whose occupation of the macroscopic state is associated with an energy, the folding free energy of the protein. In this language, a fold's designability is correlated with its sequence entropy (2).Recently, we explored the extension of the concept of equilibrium in sequence space from protein cores to the functional surface residues of proteins, specifically ligand binding sites and enzyme active sites (3). The findings of this work strongly suggested that the enzyme-substrate binding affinity, subject to constraints on the total protein energy and conformati...

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“…Biological applications have focused principally on the modeling of enzymatic catalysis and active-site chemistry (79)(80)(81)(82)(83)(84)(85)(86)(87)(88), although interesting investigations of other phenomena, such as cooperativity in backbone hydrogen bonding (89) and modeling of ␤-sheet formation propensities by a periodic DFT calculation (90), have been performed recently. The use of QM͞MM methods permits incorporation of the full protein environment, which is crucial in an important subset of cases, as has recently been shown in cytochrome P450 (79), for example.…”

Section: Applicationsmentioning

confidence: 99%

Ab initioquantum chemistry: Methodology and applications

Friesner

2005

Proc. Natl. Acad. Sci. U.S.A.

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304

230

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This Perspective provides an overview of state-of-the-art ab initio quantum chemical methodology and applications. The methods that are discussed include coupled cluster theory, localized second-order Moller-Plesset perturbation theory, multireference perturbation approaches, and density functional theory. The accuracy of each approach for key chemical properties is summarized, and the computational performance is analyzed, emphasizing significant advances in algorithms and implementation over the past decade. Incorporation of a condensed-phase environment by means of mixed quantum mechanical͞molecular mechanics or self-consistent reaction field techniques, is presented. A wide range of illustrative applications, focusing on materials science and biology, are discussed briefly.coupled cluster ͉ density functional theory ͉ second-order Modler-Plesser perturbation theory ͉ mixed quantum͞molecular mechanics O ver the past three decades, ab initio quantum chemistry has become an essential tool in the study of atoms and molecules and, increasingly, in modeling complex systems such as those arising in biology and materials science. The underlying core technology is computational solution of the electronic Schrodinger equation; given the positions of a collection of atomic nuclei, and the total number of electrons in the system, calculate the electronic energy, electron density, and other properties by means of a well defined, automated approximation (a ''model chemistry''). The ability to obtain ''goodenough'' solutions to the electronic Schrodinger equation for systems containing tens, or even hundreds, of atoms has revolutionized the ability of theoretical chemistry to address important problems in a wide range of disciplines; the Nobel Prize awarded to John Pople and Walter Kohn in 1998 is a reflection of this observation.In its exact form, the electronic Schrodinger equation is a many-body problem, whose computational complexity grows exponentially with the number of electrons, and hence, a brute force solution is intractable. Hartree-Fock theory, a mean field approach, produces reasonable results for many properties but is incapable of providing a robust description of reactive chemical events in which electron correlation has a major role. Thus, a key problem has been the development of treatments of electron correlation that exhibit a tractable scaling in computational effort with the size of the system. The considerable progress that has been made along these lines is outlined below.Given a well defined theoretical framework of approximation, the next requirement is efficient computational implementation. Considerable sophistication is required to achieve acceptable accuracy and efficiency; the leading quantum chemistry programs are millions of lines of computer code, and mathematical algorithms to reduce formal scaling of computational effort with system size have an increasingly crucial role in meeting the challenge of handling complex system relevant to practical applications. Below, the most important comp...

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Mixed Quantum Mechanical/Molecular Mechanical (QM/MM) Study of the Deacylation Reaction in a Penicillin Binding Protein (PBP) versus in a Class C β-Lactamase

Cited by 76 publications

References 85 publications

The Deacylation Mechanism of AmpC β-Lactamase at Ultrahigh Resolution

The Deacylation Mechanism of AmpC β-Lactamase at Ultrahigh Resolution

Sequence optimization and designability of enzyme active sites

Ab initioquantum chemistry: Methodology and applications

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