scholarly journals Uncovering the Binding Mode of γ-Secretase Inhibitors

2019 ◽  
Author(s):  
M. Hitzenberger ◽  
M. Zacharias
Keyword(s):  
Experimental Data ◽  
Transition State ◽  
Amyloid Precursor Protein ◽  
Active Site ◽  
Structural Model ◽  
Ph Value ◽  
Binding Mode ◽  
Enzyme Substrate ◽  
Transition State Geometry ◽  
The Stability

AbstractKnowledge of how transition state inhibitors bind to γ-secretase is of major importance for the design of new Alzheimer’s disease therapies. Based on the known structure of γ-secretase in complex with a fragment of the amyloid precursor protein we have generated a structural model of γ-secretase in complex with the effective L-685,458 transition state inhibitor. The predicted binding mode is in excellent agreement with experimental data, mimicking all enzyme-substrate interactions at the active site and forming the relevant transition state geometry with the active site aspartate residues. In addition, we found that the stability of the complex is very likely also sensitive to the pH value. Comparative simulations on the binding of L-685,458 and the epimer L682,679 allowed us to explain the strongly reduced affinity of the epimer for γ-secretase. The structural model could form a valuable basis for the design of new or modified γ-secretase inhibitors.

THEORY OF THE TRANSIENT PHASE IN AN ENZYME SYSTEM INVOLVING TWO ENZYME-SUBSTRATE COMPLEXES: THE CASE OF THE FORMATION OF PRODUCTS FROM THE FIRST COMPLEX

1959 ◽  
Vol 37 (4) ◽  
pp. 737-743 ◽  
Author(s):  
Ludovic Ouellet ◽  
James A. Stewart
Keyword(s):  
Experimental Data ◽  
Steady State ◽  
Active Site ◽  
Substrate Concentration ◽  
Enzyme System ◽  
Kinetic Scheme ◽  
Enzyme Concentration ◽  
Theoretical Treatment ◽  
Transient Phase ◽  
Enzyme Substrate

A theoretical treatment is worked out for the kinetic scheme[Formula: see text]in which the concentration of P1 is followed. The steady-state and transient phase equations are obtained subject to the condition that the substrate concentration is greatly in excess of the enzyme concentration. The conditions under which evidence in favor of this mechanism can be obtained from experimental data are discussed. Under certain conditions, the weight of the enzyme corresponding to one active site can be determined. Methods for the evaluation of the different constants are described.

Human cholinesterases

2020 ◽  
pp. 63-120
Author(s):  
Sergey Varfolomeev ◽  
Bella Grigorenko ◽  
Sofya Lushchekina ◽  
Alexander Nemuchin
Keyword(s):  
Active Site ◽  
The Body ◽  
Polymorphic Modification ◽  
Substrate Complex ◽  
Enzyme Substrate ◽  
Mechanism Of Hydrolysis ◽  
The Central Nervous System ◽  
The Stability ◽  
Hydrolysis Of ◽  
Enzyme Reactivity

The work is devoted to modeling the elementary stages of the hydrolysis reaction in the active site of enzymes belonging to the class of cholinesterases — acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). The study allowed to describe at the molecular level the effect of the polymorphic modification of BChE, causing serious physiolog ical consequences. Cholinesterase plays a crucial role in the human body. AChE is one of the key enzymes of the central nervous system, and BChE performs protective functions in the body. According to the results of calculations using the combined method of quantum and molecular mechanics (KM/MM), the mechanism of the hydrolysis of the native acetylcholine substrate in the AChE active center was detailed. For a series of ester substrates, a method for estimation of dependence of the enzyme reactivity on the structure of the substrate has been developed. The mechanism of hydrolysis of the muscle relaxant of succininylcholine BChE and the effect of the Asp70Gly polymorph on it were studied. Using various computer simulation methods, the stability of the enzyme-substrate complex of two enzyme variants with succinylcholine was studied.

scholarly journals Construction of a Full-Atomic Mechanistic Model of Human Apurinic/Apyrimidinic Endonuclease APE1 for Virtual Screening of Novel Inhibitors

Acta Naturae ◽  
2012 ◽  
Vol 4 (2) ◽  
pp. 80-86
Author(s):  
I. G. Khaliullin ◽  
D. K. Nilov ◽  
I. V. Shapovalova ◽  
V. K. Švedas
Keyword(s):  
Experimental Data ◽  
Active Site ◽  
Molecular Model ◽  
Mechanistic Model ◽  
Proton Acceptor ◽  
Repair System ◽  
Amino Acid Residues ◽  
Enzyme Substrate ◽  
Oncological Diseases ◽  
Important Enzyme

A full-atomic molecular model of human apurinic/apyrimidinic endonuclease APE1, an important enzyme in the DNA repair system, has been constructed. The research consisted of hybrid quantum mechanics/ molecular mechanics modeling of the enzyme-substrate interactions, as well as calculations of the ionization states of the amino acid residues of the active site of the enzyme. The choice of the APE1 mechanism with an Asp210 residue as a proton acceptor was validated by means of a generalization of modeling and experimental data. Interactions were revealed in the active site that are of greatest significance for binding the substrate and potential APE1 inhibitors (potential co-drugs of interest in the chemo- and radiotherapy of oncological diseases).

scholarly journals Evolutionary repurposing of a sulfatase: A new Michaelis complex leads to efficient transition state charge offset

2018 ◽  
Vol 115 (31) ◽  
pp. E7293-E7302 ◽  
Author(s):  
Charlotte M. Miton ◽  
Stefanie Jonas ◽  
Gerhard Fischer ◽  
Fernanda Duarte ◽  
Mark F. Mohamed ◽  
...  
Keyword(s):  
Molecular Recognition ◽  
Transition State ◽  
Active Site ◽  
High Reactivity ◽  
Fine Tuning ◽  
Binding Modes ◽  
Evolutionary Trajectory ◽  
Multiple Substrates ◽  
Enzyme Substrate ◽  
Multiple Substrate

The recruitment and evolutionary optimization of promiscuous enzymes is key to the rapid adaptation of organisms to changing environments. Our understanding of the precise mechanisms underlying enzyme repurposing is, however, limited: What are the active-site features that enable the molecular recognition of multiple substrates with contrasting catalytic requirements? To gain insights into the molecular determinants of adaptation in promiscuous enzymes, we performed the laboratory evolution of an arylsulfatase to improve its initially weak phenylphosphonate hydrolase activity. The evolutionary trajectory led to a 100,000-fold enhancement of phenylphosphonate hydrolysis, while the native sulfate and promiscuous phosphate mono- and diester hydrolyses were only marginally affected (≤50-fold). Structural, kinetic, and in silico characterizations of the evolutionary intermediates revealed that two key mutations, T50A and M72V, locally reshaped the active site, improving access to the catalytic machinery for the phosphonate. Measured transition state (TS) charge changes along the trajectory suggest the creation of a new Michaelis complex (E•S, enzyme–substrate), with enhanced leaving group stabilization in the TS for the promiscuous phosphonate (βleavinggroup from −1.08 to −0.42). Rather than altering the catalytic machinery, evolutionary repurposing was achieved by fine-tuning the molecular recognition of the phosphonate in the Michaelis complex, and by extension, also in the TS. This molecular scenario constitutes a mechanistic alternative to adaptation solely based on enzyme flexibility and conformational selection. Instead, rapid functional transitions between distinct chemical reactions rely on the high reactivity of permissive active-site architectures that allow multiple substrate binding modes.

Human cholinesterases

2020 ◽  
pp. 69-126
Author(s):  
Sergey Varfolomeev ◽  
Bella Grigorenko ◽  
Sofya Lushchekina ◽  
Patrick Masson ◽  
Galina Mahaeva ◽  
...  
Keyword(s):  
Active Site ◽  
The Body ◽  
Polymorphic Modification ◽  
Substrate Complex ◽  
Enzyme Substrate ◽  
Mechanism Of Hydrolysis ◽  
The Central Nervous System ◽  
The Stability ◽  
Hydrolysis Of ◽  
Enzyme Reactivity

The work is devoted to modeling the elementary stages of the hydrolysis reaction in the active site of enzymes belonging to the class of cholinesterases — acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). The study allowed to describe at the molecular level the effect of the polymorphic modification of BChE, causing serious physiolog ical consequences. Cholinesterase plays a crucial role in the human body. AChE is one of the key enzymes of the central nervous system, and BChE performs protective functions in the body. According to the results of calculations using the combined method of quantum and molecular mechanics (KM/MM), the mechanism of the hydrolysis of the native acetylcholine substrate in the AChE active center was detailed. For a series of ester substrates, a method for estimation of dependence of the enzyme reactivity on the structure of the substrate has been developed. The mechanism of hydrolysis of the muscle relaxant of succininylcholine BChE and the effect of the Asp70Gly polymorph on it were studied. Using various computer simulation methods, the stability of the enzyme-substrate complex of two enzyme variants with succinylcholine was studied.

scholarly journals Biochemical characterization of the chondroitinase ABC I active site

2005 ◽  
Vol 390 (2) ◽  
pp. 395-405 ◽  
Author(s):  
Vikas Prabhakar ◽  
Rahul Raman ◽  
Ishan Capila ◽  
Carlos J. Bosques ◽  
Kevin Pojasek ◽  
...  
Keyword(s):  
Active Site ◽  
Structural Model ◽  
Catalytic Mechanism ◽  
Biochemical Characterization ◽  
Chondroitin Sulphate ◽  
Critical Role ◽  
Site Directed Mutagenesis ◽  
Chondroitinase Abc ◽  
Enzyme Substrate

cABC I (chondroitinase ABC I) from Proteus vulgaris is a GalAG (galactosaminoglycan) depolymerizing lyase that cleaves its substrates at the glycosidic bond via β-elimination. cABC I cleaves a particularly broad range of GalAG substrates, including CS (chondroitin sulphate), DS (dermatan sulphate) and hyaluronic acid. We recently cloned and recombinantly expressed cABC I in Escherichia coli, and completed a preliminary biochemical characterization of the enzyme. In the present study, we have coupled site-directed mutagenesis of the recombinant cABC I with a structural model of the enzyme–substrate complex in order to investigate in detail the roles of active site amino acids in the catalytic action of the enzyme. The putative catalytic residues His-501, Tyr-508, Arg-560 and Glu-653 were probed systematically via mutagenesis. Assessment of these mutants in kinetic and end-point assays provided direct evidence on the catalytic roles of these active-site residues. The crystal structure of the native enzyme provided a framework for molecular docking of representative CS and DS substrates. This enabled us to construct recombinant enzyme–substrate structural complexes. These studies together provided structural insights into the effects of the mutations on the catalytic mechanism of cABC I and the differences in its processing of CS and DS substrates. All His-501 mutants were essentially inactive and thereby implicating this amino acid to play the critical role of proton abstraction during catalysis. The kinetic data for Glu-653 mutants indicated that it is involved in a hydrogen bonding network in the active site. The proximity of Tyr-508 to the glycosidic oxygen of the substrate at the site of cleavage suggested its potential role in protonating the leaving group. Arg-560 was proximal to the uronic acid C-5 proton, suggesting its possible role in the stabilization of the carbanion intermediate formed during catalysis.

scholarly journals Construction of a Full-Atomic Mechanistic Model of Human Apurinic/Apyrimidinic Endonuclease APE1 for Virtual Screening of Novel Inhibitors

Acta Naturae ◽  
2012 ◽  
Vol 4 (2) ◽  
pp. 80-86 ◽  
Author(s):  
I. G. Khaliullin ◽  
D. K. Nilov ◽  
I. V. Shapovalova ◽  
V. K. Švedas
Keyword(s):  
Experimental Data ◽  
Active Site ◽  
Molecular Model ◽  
Mechanistic Model ◽  
Proton Acceptor ◽  
Repair System ◽  
Amino Acid Residues ◽  
Enzyme Substrate ◽  
Oncological Diseases ◽  
Important Enzyme

A full-atomic molecular model of human apurinic/apyrimidinic endonuclease APE1, an important enzyme in the DNA repair system, has been constructed. The research consisted of hybrid quantum mechanics/ molecular mechanics modeling of the enzyme-substrate interactions, as well as calculations of the ionization states of the amino acid residues of the active site of the enzyme. The choice of the APE1 mechanism with an Asp210 residue as a proton acceptor was validated by means of a generalization of modeling and experimental data. Interactions were revealed in the active site that are of greatest significance for binding the substrate and potential APE1 inhibitors (potential co-drugs of interest in the chemo- and radiotherapy of oncological diseases).

Memapsin 2, a drug target for Alzheimer's disease

2003 ◽  
Vol 70 ◽  
pp. 213-220 ◽  
Author(s):  
Gerald Koelsch ◽  
Robert T. Turner ◽  
Lin Hong ◽  
Arun K. Ghosh ◽  
Jordan Tang
Keyword(s):  
Crystal Structure ◽  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Transition State ◽  
Amyloid Precursor Protein ◽  
Therapeutic Target ◽  
Drug Target ◽  
Aspartic Protease ◽  
Precursor Protein ◽  
Memapsin 2

Mempasin 2, a ϐ-secretase, is the membrane-anchored aspartic protease that initiates the cleavage of amyloid precursor protein leading to the production of ϐ-amyloid and the onset of Alzheimer's disease. Thus memapsin 2 is a major therapeutic target for the development of inhibitor drugs for the disease. Many biochemical tools, such as the specificity and crystal structure, have been established and have led to the design of potent and relatively small transition-state inhibitors. Although developing a clinically viable mempasin 2 inhibitor remains challenging, progress to date renders hope that memapsin 2 inhibitors may ultimately be useful for therapeutic reduction of ϐ-amyloid.

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