Kinetic measurement of the interaction between a lysozyme and its immobilized substrate analogue by means of surface plasmon resonance

Tadashi Ueda, Miyako Tsurumaru, Taiji Imoto

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

A method for evaluating the association and dissociation rate constants of interaction between a lysozyme and its substrate analogue, an immobilized p-aminophenyl-tri-N-acetyl-β-chitotrioside, by means of surface plasmon resonance has been developed. Site-specific immobilization of p-aminophenyl-tri-N-acetyl-β-chitotrioside, which is a product of p-nitrophenyl-tri-N-acetyl-β-chitotrioside, on carboxymethyldextran linked to the surface of the cuvette of the instrument, IAsys, was carried out by catalysis with EDC/NHS. The kinetic parameters of the interaction between hen or human lysozyme and the immobilized substrate analogue indicated that a larger dissociation constant of the human lysozyme-immobilized substrate analogue complex depended on a smaller association rate constant. The kinetic parameters of the interaction between the immobilized substrate analogue and a mutant hen lysozyme, in which Arg14 and His15 are deleted, with higher activity than the wild type hen lysozyme were measured. It was suggested that the higher activity of the mutant lysozyme was due to faster removal of the substrate from the active site cleft and/or the formation of a stabler and better complex as to hydrolysis.

Original languageEnglish
Pages (from-to)712-716
Number of pages5
JournalJournal of Biochemistry
Volume124
Issue number4
DOIs
Publication statusPublished - Jan 1 1998

Fingerprint

Surface Plasmon Resonance
Surface plasmon resonance
Muramidase
Kinetics
Substrates
Kinetic parameters
Rate constants
Association reactions
Catalysis
Immobilization
Hydrolysis
Catalytic Domain

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology

Cite this

Kinetic measurement of the interaction between a lysozyme and its immobilized substrate analogue by means of surface plasmon resonance. / Ueda, Tadashi; Tsurumaru, Miyako; Imoto, Taiji.

In: Journal of Biochemistry, Vol. 124, No. 4, 01.01.1998, p. 712-716.

Research output: Contribution to journalArticle

@article{c40831b7524947df9d3418fcc27bd9c8,
title = "Kinetic measurement of the interaction between a lysozyme and its immobilized substrate analogue by means of surface plasmon resonance",
abstract = "A method for evaluating the association and dissociation rate constants of interaction between a lysozyme and its substrate analogue, an immobilized p-aminophenyl-tri-N-acetyl-β-chitotrioside, by means of surface plasmon resonance has been developed. Site-specific immobilization of p-aminophenyl-tri-N-acetyl-β-chitotrioside, which is a product of p-nitrophenyl-tri-N-acetyl-β-chitotrioside, on carboxymethyldextran linked to the surface of the cuvette of the instrument, IAsys, was carried out by catalysis with EDC/NHS. The kinetic parameters of the interaction between hen or human lysozyme and the immobilized substrate analogue indicated that a larger dissociation constant of the human lysozyme-immobilized substrate analogue complex depended on a smaller association rate constant. The kinetic parameters of the interaction between the immobilized substrate analogue and a mutant hen lysozyme, in which Arg14 and His15 are deleted, with higher activity than the wild type hen lysozyme were measured. It was suggested that the higher activity of the mutant lysozyme was due to faster removal of the substrate from the active site cleft and/or the formation of a stabler and better complex as to hydrolysis.",
author = "Tadashi Ueda and Miyako Tsurumaru and Taiji Imoto",
year = "1998",
month = "1",
day = "1",
doi = "10.1093/oxfordjournals.jbchem.a022171",
language = "English",
volume = "124",
pages = "712--716",
journal = "Journal of Biochemistry",
issn = "0021-924X",
publisher = "Oxford University Press",
number = "4",

}

TY - JOUR

T1 - Kinetic measurement of the interaction between a lysozyme and its immobilized substrate analogue by means of surface plasmon resonance

AU - Ueda, Tadashi

AU - Tsurumaru, Miyako

AU - Imoto, Taiji

PY - 1998/1/1

Y1 - 1998/1/1

N2 - A method for evaluating the association and dissociation rate constants of interaction between a lysozyme and its substrate analogue, an immobilized p-aminophenyl-tri-N-acetyl-β-chitotrioside, by means of surface plasmon resonance has been developed. Site-specific immobilization of p-aminophenyl-tri-N-acetyl-β-chitotrioside, which is a product of p-nitrophenyl-tri-N-acetyl-β-chitotrioside, on carboxymethyldextran linked to the surface of the cuvette of the instrument, IAsys, was carried out by catalysis with EDC/NHS. The kinetic parameters of the interaction between hen or human lysozyme and the immobilized substrate analogue indicated that a larger dissociation constant of the human lysozyme-immobilized substrate analogue complex depended on a smaller association rate constant. The kinetic parameters of the interaction between the immobilized substrate analogue and a mutant hen lysozyme, in which Arg14 and His15 are deleted, with higher activity than the wild type hen lysozyme were measured. It was suggested that the higher activity of the mutant lysozyme was due to faster removal of the substrate from the active site cleft and/or the formation of a stabler and better complex as to hydrolysis.

AB - A method for evaluating the association and dissociation rate constants of interaction between a lysozyme and its substrate analogue, an immobilized p-aminophenyl-tri-N-acetyl-β-chitotrioside, by means of surface plasmon resonance has been developed. Site-specific immobilization of p-aminophenyl-tri-N-acetyl-β-chitotrioside, which is a product of p-nitrophenyl-tri-N-acetyl-β-chitotrioside, on carboxymethyldextran linked to the surface of the cuvette of the instrument, IAsys, was carried out by catalysis with EDC/NHS. The kinetic parameters of the interaction between hen or human lysozyme and the immobilized substrate analogue indicated that a larger dissociation constant of the human lysozyme-immobilized substrate analogue complex depended on a smaller association rate constant. The kinetic parameters of the interaction between the immobilized substrate analogue and a mutant hen lysozyme, in which Arg14 and His15 are deleted, with higher activity than the wild type hen lysozyme were measured. It was suggested that the higher activity of the mutant lysozyme was due to faster removal of the substrate from the active site cleft and/or the formation of a stabler and better complex as to hydrolysis.

UR - http://www.scopus.com/inward/record.url?scp=0031790675&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0031790675&partnerID=8YFLogxK

U2 - 10.1093/oxfordjournals.jbchem.a022171

DO - 10.1093/oxfordjournals.jbchem.a022171

M3 - Article

C2 - 9756615

AN - SCOPUS:0031790675

VL - 124

SP - 712

EP - 716

JO - Journal of Biochemistry

JF - Journal of Biochemistry

SN - 0021-924X

IS - 4

ER -