Effects of viscosity and osmotic stress on the reaction of human butyrylcholinesterase with cresyl saligenin phosphate, a toxicant related to aerotoxic syndrome: Kinetic and molecular dynamics studies

Patrick Masson, Sofya Lushchekina, Lawrence M Schopfer, Oksana Lockridge

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28 Citations (Scopus)

Abstract

CSP (cresyl saligenin phosphate) is an irreversible inhibitor of human BChE (butyrylcholinesterase) that has been involved in the aerotoxic syndrome. Inhibition under pseudo-first-order conditions is biphasic, reflecting a slow equilibrium between two enzyme states E and E′. The elementary constants for CSP inhibition of wild-type BChE and D70G mutant were determined by studying the dependence of inhibition kinetics on viscosity and osmotic pressure. Glycerol and sucrose were used as viscosogens. Phosphorylation by CSP is sensitive to viscosity and is thus strongly diffusion-controlled (k on≈108 M-1 · min-1). Bimolecular rate constants (ki) are about equal to kon values, making CSP one of the fastest inhibitors of BChE. Sucrose caused osmotic stress because it is excluded from the active-site gorge. This depleted the active-site gorge of water. Osmotic activation volumes, determined from the dependence of ki on osmotic pressure, showed that water in the gorge of the D70G mutant is more easily depleted than that in wild-type BChE. This demonstrates the importance of the peripheral site residue Asp70 in controlling the active-site gorge hydration. MD simulations provided new evidence for differences in the motion of water within the gorge of wild-type and D70G enzymes. The effect of viscosogens/osmolytes provided information on the slow equilibrium E⇌E′, indicating that alteration in hydration of a key catalytic residue shifts the equilibrium towards E′. MD simulations showed that glycerol molecules that substitute for water molecules in the enzyme active-site gorge induce a conformational change in the catalytic triad residue His438, leading to the less reactive form E′.

Original languageEnglish (US)
Pages (from-to)387-399
Number of pages13
JournalBiochemical Journal
Volume454
Issue number3
DOIs
StatePublished - Sep 15 2013

Fingerprint

Butyrylcholinesterase
Osmotic Pressure
Molecular Dynamics Simulation
Viscosity
Molecular dynamics
Catalytic Domain
Phosphates
Kinetics
Water
Hydration
Glycerol
Sucrose
Enzymes
Phosphorylation
Molecules
Rate constants
Chemical activation
salicyl alcohol

Keywords

  • Aerotoxic syndrome
  • Butyrylcholinesterase (BChE)
  • Cresyl saligenin phosphate (CSP)
  • Diffusion control
  • Organophosphate
  • Osmotic stress
  • Viscosity

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

@article{57e164d4275544a695d149ff6a79221c,
title = "Effects of viscosity and osmotic stress on the reaction of human butyrylcholinesterase with cresyl saligenin phosphate, a toxicant related to aerotoxic syndrome: Kinetic and molecular dynamics studies",
abstract = "CSP (cresyl saligenin phosphate) is an irreversible inhibitor of human BChE (butyrylcholinesterase) that has been involved in the aerotoxic syndrome. Inhibition under pseudo-first-order conditions is biphasic, reflecting a slow equilibrium between two enzyme states E and E′. The elementary constants for CSP inhibition of wild-type BChE and D70G mutant were determined by studying the dependence of inhibition kinetics on viscosity and osmotic pressure. Glycerol and sucrose were used as viscosogens. Phosphorylation by CSP is sensitive to viscosity and is thus strongly diffusion-controlled (k on≈108 M-1 · min-1). Bimolecular rate constants (ki) are about equal to kon values, making CSP one of the fastest inhibitors of BChE. Sucrose caused osmotic stress because it is excluded from the active-site gorge. This depleted the active-site gorge of water. Osmotic activation volumes, determined from the dependence of ki on osmotic pressure, showed that water in the gorge of the D70G mutant is more easily depleted than that in wild-type BChE. This demonstrates the importance of the peripheral site residue Asp70 in controlling the active-site gorge hydration. MD simulations provided new evidence for differences in the motion of water within the gorge of wild-type and D70G enzymes. The effect of viscosogens/osmolytes provided information on the slow equilibrium E⇌E′, indicating that alteration in hydration of a key catalytic residue shifts the equilibrium towards E′. MD simulations showed that glycerol molecules that substitute for water molecules in the enzyme active-site gorge induce a conformational change in the catalytic triad residue His438, leading to the less reactive form E′.",
keywords = "Aerotoxic syndrome, Butyrylcholinesterase (BChE), Cresyl saligenin phosphate (CSP), Diffusion control, Organophosphate, Osmotic stress, Viscosity",
author = "Patrick Masson and Sofya Lushchekina and Schopfer, {Lawrence M} and Oksana Lockridge",
year = "2013",
month = "9",
day = "15",
doi = "10.1042/BJ20130389",
language = "English (US)",
volume = "454",
pages = "387--399",
journal = "Biochemical Journal",
issn = "0264-6021",
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T1 - Effects of viscosity and osmotic stress on the reaction of human butyrylcholinesterase with cresyl saligenin phosphate, a toxicant related to aerotoxic syndrome

T2 - Kinetic and molecular dynamics studies

AU - Masson, Patrick

AU - Lushchekina, Sofya

AU - Schopfer, Lawrence M

AU - Lockridge, Oksana

PY - 2013/9/15

Y1 - 2013/9/15

N2 - CSP (cresyl saligenin phosphate) is an irreversible inhibitor of human BChE (butyrylcholinesterase) that has been involved in the aerotoxic syndrome. Inhibition under pseudo-first-order conditions is biphasic, reflecting a slow equilibrium between two enzyme states E and E′. The elementary constants for CSP inhibition of wild-type BChE and D70G mutant were determined by studying the dependence of inhibition kinetics on viscosity and osmotic pressure. Glycerol and sucrose were used as viscosogens. Phosphorylation by CSP is sensitive to viscosity and is thus strongly diffusion-controlled (k on≈108 M-1 · min-1). Bimolecular rate constants (ki) are about equal to kon values, making CSP one of the fastest inhibitors of BChE. Sucrose caused osmotic stress because it is excluded from the active-site gorge. This depleted the active-site gorge of water. Osmotic activation volumes, determined from the dependence of ki on osmotic pressure, showed that water in the gorge of the D70G mutant is more easily depleted than that in wild-type BChE. This demonstrates the importance of the peripheral site residue Asp70 in controlling the active-site gorge hydration. MD simulations provided new evidence for differences in the motion of water within the gorge of wild-type and D70G enzymes. The effect of viscosogens/osmolytes provided information on the slow equilibrium E⇌E′, indicating that alteration in hydration of a key catalytic residue shifts the equilibrium towards E′. MD simulations showed that glycerol molecules that substitute for water molecules in the enzyme active-site gorge induce a conformational change in the catalytic triad residue His438, leading to the less reactive form E′.

AB - CSP (cresyl saligenin phosphate) is an irreversible inhibitor of human BChE (butyrylcholinesterase) that has been involved in the aerotoxic syndrome. Inhibition under pseudo-first-order conditions is biphasic, reflecting a slow equilibrium between two enzyme states E and E′. The elementary constants for CSP inhibition of wild-type BChE and D70G mutant were determined by studying the dependence of inhibition kinetics on viscosity and osmotic pressure. Glycerol and sucrose were used as viscosogens. Phosphorylation by CSP is sensitive to viscosity and is thus strongly diffusion-controlled (k on≈108 M-1 · min-1). Bimolecular rate constants (ki) are about equal to kon values, making CSP one of the fastest inhibitors of BChE. Sucrose caused osmotic stress because it is excluded from the active-site gorge. This depleted the active-site gorge of water. Osmotic activation volumes, determined from the dependence of ki on osmotic pressure, showed that water in the gorge of the D70G mutant is more easily depleted than that in wild-type BChE. This demonstrates the importance of the peripheral site residue Asp70 in controlling the active-site gorge hydration. MD simulations provided new evidence for differences in the motion of water within the gorge of wild-type and D70G enzymes. The effect of viscosogens/osmolytes provided information on the slow equilibrium E⇌E′, indicating that alteration in hydration of a key catalytic residue shifts the equilibrium towards E′. MD simulations showed that glycerol molecules that substitute for water molecules in the enzyme active-site gorge induce a conformational change in the catalytic triad residue His438, leading to the less reactive form E′.

KW - Aerotoxic syndrome

KW - Butyrylcholinesterase (BChE)

KW - Cresyl saligenin phosphate (CSP)

KW - Diffusion control

KW - Organophosphate

KW - Osmotic stress

KW - Viscosity

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U2 - 10.1042/BJ20130389

DO - 10.1042/BJ20130389

M3 - Article

C2 - 23782236

AN - SCOPUS:84883378115

VL - 454

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JO - Biochemical Journal

JF - Biochemical Journal

SN - 0264-6021

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