Polyionic complexes of butyrylcholinesterase and poly-L-lysine-g-poly(ethylene glycol)

Comparative kinetics of catalysis and inhibition and in vitro inactivation by proteases and heat

Kirstin Hester, Jing Liu, Nicholas Flynn, Lester G. Sultatos, Liyi Geng, William Stephen Brimijoin, Joshua D. Ramsey, Steven Hartson, Ashish Ranjan, Carey Pope

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

We previously reported that recombinant human butyrylcholinesterase (rhBChE) complexed with a series of copolymers of poly-L-lysine (PLL) with grafted (polyethylene) glycol (PEG) (i.e., PLL-g-PEG) showed reduced catalytic activity but relatively similar concentration-dependent inactivation of the organophosphorus inhibitor paraoxon. Herein, we compared the kinetics of catalysis (using butyrylthiocholine as the substrate) and inhibition (using four different inhibitors) of free and copolymer-complexed rhBChE. Using scanning electron microscopy, polyionic complexes of rhBChE with three different PLL-g-PEG copolymers (based on PLL size) appeared as spheroid-shaped particles with relatively similar particle sizes (median diameter = 35 nm). Relatively similar particle sizes were also noted using dynamic light scattering (mean = 26–35 nm). The three copolymer-complexed enzymes exhibited reduced kcat (30–33% reduction), but no significant changes in Km. Inhibitory potency (as reflected by the bimolecular rate constant, ki) was similar among the free and copolymer-complexed enzymes when paraoxon was the inhibitor, whereas statistically significant reductions in ki (16–60%) were noted with the other inhibitors. Sensitivity to inactivation by proteases and heat was also compared. Copolymer-complexed enzymes showed lesser time-dependent inactivation by the proteases trypsin and pronase and by heat compared to the free enzyme. Understanding the unique properties of PLL-g-PEG-BChE complexes may lead to enhanced approaches for use of BChE and other protein bioscavengers.

Original languageEnglish (US)
Pages (from-to)86-94
Number of pages9
JournalChemico-Biological Interactions
Volume275
DOIs
StatePublished - Sep 25 2017

Fingerprint

Butyrylcholinesterase
Ethylene Glycol
Catalysis
Polyethylene glycols
Lysine
Peptide Hydrolases
Copolymers
Hot Temperature
Kinetics
Paraoxon
Enzymes
Particle Size
Butyrylthiocholine
Particle size
Pronase
Electron Scanning Microscopy
Trypsin
Dynamic light scattering
Rate constants
Catalyst activity

Keywords

  • Bioscavenger
  • Copolymer-BChE complex
  • Organophosphates
  • Recombinant BChE

ASJC Scopus subject areas

  • Toxicology

Cite this

Polyionic complexes of butyrylcholinesterase and poly-L-lysine-g-poly(ethylene glycol) : Comparative kinetics of catalysis and inhibition and in vitro inactivation by proteases and heat. / Hester, Kirstin; Liu, Jing; Flynn, Nicholas; Sultatos, Lester G.; Geng, Liyi; Brimijoin, William Stephen; Ramsey, Joshua D.; Hartson, Steven; Ranjan, Ashish; Pope, Carey.

In: Chemico-Biological Interactions, Vol. 275, 25.09.2017, p. 86-94.

Research output: Contribution to journalArticle

Hester, Kirstin ; Liu, Jing ; Flynn, Nicholas ; Sultatos, Lester G. ; Geng, Liyi ; Brimijoin, William Stephen ; Ramsey, Joshua D. ; Hartson, Steven ; Ranjan, Ashish ; Pope, Carey. / Polyionic complexes of butyrylcholinesterase and poly-L-lysine-g-poly(ethylene glycol) : Comparative kinetics of catalysis and inhibition and in vitro inactivation by proteases and heat. In: Chemico-Biological Interactions. 2017 ; Vol. 275. pp. 86-94.
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abstract = "We previously reported that recombinant human butyrylcholinesterase (rhBChE) complexed with a series of copolymers of poly-L-lysine (PLL) with grafted (polyethylene) glycol (PEG) (i.e., PLL-g-PEG) showed reduced catalytic activity but relatively similar concentration-dependent inactivation of the organophosphorus inhibitor paraoxon. Herein, we compared the kinetics of catalysis (using butyrylthiocholine as the substrate) and inhibition (using four different inhibitors) of free and copolymer-complexed rhBChE. Using scanning electron microscopy, polyionic complexes of rhBChE with three different PLL-g-PEG copolymers (based on PLL size) appeared as spheroid-shaped particles with relatively similar particle sizes (median diameter = 35 nm). Relatively similar particle sizes were also noted using dynamic light scattering (mean = 26–35 nm). The three copolymer-complexed enzymes exhibited reduced kcat (30–33{\%} reduction), but no significant changes in Km. Inhibitory potency (as reflected by the bimolecular rate constant, ki) was similar among the free and copolymer-complexed enzymes when paraoxon was the inhibitor, whereas statistically significant reductions in ki (16–60{\%}) were noted with the other inhibitors. Sensitivity to inactivation by proteases and heat was also compared. Copolymer-complexed enzymes showed lesser time-dependent inactivation by the proteases trypsin and pronase and by heat compared to the free enzyme. Understanding the unique properties of PLL-g-PEG-BChE complexes may lead to enhanced approaches for use of BChE and other protein bioscavengers.",
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T1 - Polyionic complexes of butyrylcholinesterase and poly-L-lysine-g-poly(ethylene glycol)

T2 - Comparative kinetics of catalysis and inhibition and in vitro inactivation by proteases and heat

AU - Hester, Kirstin

AU - Liu, Jing

AU - Flynn, Nicholas

AU - Sultatos, Lester G.

AU - Geng, Liyi

AU - Brimijoin, William Stephen

AU - Ramsey, Joshua D.

AU - Hartson, Steven

AU - Ranjan, Ashish

AU - Pope, Carey

PY - 2017/9/25

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N2 - We previously reported that recombinant human butyrylcholinesterase (rhBChE) complexed with a series of copolymers of poly-L-lysine (PLL) with grafted (polyethylene) glycol (PEG) (i.e., PLL-g-PEG) showed reduced catalytic activity but relatively similar concentration-dependent inactivation of the organophosphorus inhibitor paraoxon. Herein, we compared the kinetics of catalysis (using butyrylthiocholine as the substrate) and inhibition (using four different inhibitors) of free and copolymer-complexed rhBChE. Using scanning electron microscopy, polyionic complexes of rhBChE with three different PLL-g-PEG copolymers (based on PLL size) appeared as spheroid-shaped particles with relatively similar particle sizes (median diameter = 35 nm). Relatively similar particle sizes were also noted using dynamic light scattering (mean = 26–35 nm). The three copolymer-complexed enzymes exhibited reduced kcat (30–33% reduction), but no significant changes in Km. Inhibitory potency (as reflected by the bimolecular rate constant, ki) was similar among the free and copolymer-complexed enzymes when paraoxon was the inhibitor, whereas statistically significant reductions in ki (16–60%) were noted with the other inhibitors. Sensitivity to inactivation by proteases and heat was also compared. Copolymer-complexed enzymes showed lesser time-dependent inactivation by the proteases trypsin and pronase and by heat compared to the free enzyme. Understanding the unique properties of PLL-g-PEG-BChE complexes may lead to enhanced approaches for use of BChE and other protein bioscavengers.

AB - We previously reported that recombinant human butyrylcholinesterase (rhBChE) complexed with a series of copolymers of poly-L-lysine (PLL) with grafted (polyethylene) glycol (PEG) (i.e., PLL-g-PEG) showed reduced catalytic activity but relatively similar concentration-dependent inactivation of the organophosphorus inhibitor paraoxon. Herein, we compared the kinetics of catalysis (using butyrylthiocholine as the substrate) and inhibition (using four different inhibitors) of free and copolymer-complexed rhBChE. Using scanning electron microscopy, polyionic complexes of rhBChE with three different PLL-g-PEG copolymers (based on PLL size) appeared as spheroid-shaped particles with relatively similar particle sizes (median diameter = 35 nm). Relatively similar particle sizes were also noted using dynamic light scattering (mean = 26–35 nm). The three copolymer-complexed enzymes exhibited reduced kcat (30–33% reduction), but no significant changes in Km. Inhibitory potency (as reflected by the bimolecular rate constant, ki) was similar among the free and copolymer-complexed enzymes when paraoxon was the inhibitor, whereas statistically significant reductions in ki (16–60%) were noted with the other inhibitors. Sensitivity to inactivation by proteases and heat was also compared. Copolymer-complexed enzymes showed lesser time-dependent inactivation by the proteases trypsin and pronase and by heat compared to the free enzyme. Understanding the unique properties of PLL-g-PEG-BChE complexes may lead to enhanced approaches for use of BChE and other protein bioscavengers.

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KW - Copolymer-BChE complex

KW - Organophosphates

KW - Recombinant BChE

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