Site-specifically labeled photoprotein-thyroxine conjugates using aequorin mutants containing unique cysteine residues: Applications for binding assays (Part II)

J. C. Lewis, L. C. Cullen, Sylvia Daunert

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

The jellyfish Aequorea victoria produces a protein, aequorin, which belongs to the class of Ca2+-dependent photoproteins known for their ability to emit visible light. This property of aequorin has allowed for its as a bioluminescent label in binding assays for a variety of analytes. Due to the excellent detection limits we demonstrated in assays for small peptides using a fusion protein between the peptide of interest and the photoprotein, our next goal was to expand the range of possible analytes for producing homogeneous populations of conjugates with the aequorin label to those that were nonpeptidic in nature. Recently, we prepared and characterized four aequorin mutants containing unique cysteine residues at various positions in the polypeptide chain. In the work reported here, the four aequorin mutants were each conjugated with a maleimide-activated methyl ester derivative of thyroxine, a hormone frequently determined to evaluate thyroid function. The thyroxine-aequorin mutant conjugates were characterized in terms of the bioluminescence activities and binding properties with an anti-thyroxine monoclonal antibody for possible future employment in either heterogeneous or homogeneous binding assays for thyroxine and/or other desired analytes.

Original languageEnglish
Pages (from-to)140-145
Number of pages6
JournalBioconjugate Chemistry
Volume11
Issue number2
DOIs
StatePublished - Mar 1 2000
Externally publishedYes

Fingerprint

Luminescent Proteins
Aequorin
Thyroxine
Cysteine
Assays
Peptides
Labels
Bioluminescence
Proteins
Monoclonal antibodies
Polypeptides
Hormones
Esters
Fusion reactions
Derivatives
Limit of Detection
Thyroid Gland
Monoclonal Antibodies
Light
Population

ASJC Scopus subject areas

  • Chemistry(all)
  • Organic Chemistry
  • Clinical Biochemistry
  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry

Cite this

@article{7f9b65a9ebc147d0912cd68d9fb1bcd0,
title = "Site-specifically labeled photoprotein-thyroxine conjugates using aequorin mutants containing unique cysteine residues: Applications for binding assays (Part II)",
abstract = "The jellyfish Aequorea victoria produces a protein, aequorin, which belongs to the class of Ca2+-dependent photoproteins known for their ability to emit visible light. This property of aequorin has allowed for its as a bioluminescent label in binding assays for a variety of analytes. Due to the excellent detection limits we demonstrated in assays for small peptides using a fusion protein between the peptide of interest and the photoprotein, our next goal was to expand the range of possible analytes for producing homogeneous populations of conjugates with the aequorin label to those that were nonpeptidic in nature. Recently, we prepared and characterized four aequorin mutants containing unique cysteine residues at various positions in the polypeptide chain. In the work reported here, the four aequorin mutants were each conjugated with a maleimide-activated methyl ester derivative of thyroxine, a hormone frequently determined to evaluate thyroid function. The thyroxine-aequorin mutant conjugates were characterized in terms of the bioluminescence activities and binding properties with an anti-thyroxine monoclonal antibody for possible future employment in either heterogeneous or homogeneous binding assays for thyroxine and/or other desired analytes.",
author = "Lewis, {J. C.} and Cullen, {L. C.} and Sylvia Daunert",
year = "2000",
month = "3",
day = "1",
doi = "10.1021/bc990081s",
language = "English",
volume = "11",
pages = "140--145",
journal = "Bioconjugate Chemistry",
issn = "1043-1802",
publisher = "American Chemical Society",
number = "2",

}

TY - JOUR

T1 - Site-specifically labeled photoprotein-thyroxine conjugates using aequorin mutants containing unique cysteine residues

T2 - Applications for binding assays (Part II)

AU - Lewis, J. C.

AU - Cullen, L. C.

AU - Daunert, Sylvia

PY - 2000/3/1

Y1 - 2000/3/1

N2 - The jellyfish Aequorea victoria produces a protein, aequorin, which belongs to the class of Ca2+-dependent photoproteins known for their ability to emit visible light. This property of aequorin has allowed for its as a bioluminescent label in binding assays for a variety of analytes. Due to the excellent detection limits we demonstrated in assays for small peptides using a fusion protein between the peptide of interest and the photoprotein, our next goal was to expand the range of possible analytes for producing homogeneous populations of conjugates with the aequorin label to those that were nonpeptidic in nature. Recently, we prepared and characterized four aequorin mutants containing unique cysteine residues at various positions in the polypeptide chain. In the work reported here, the four aequorin mutants were each conjugated with a maleimide-activated methyl ester derivative of thyroxine, a hormone frequently determined to evaluate thyroid function. The thyroxine-aequorin mutant conjugates were characterized in terms of the bioluminescence activities and binding properties with an anti-thyroxine monoclonal antibody for possible future employment in either heterogeneous or homogeneous binding assays for thyroxine and/or other desired analytes.

AB - The jellyfish Aequorea victoria produces a protein, aequorin, which belongs to the class of Ca2+-dependent photoproteins known for their ability to emit visible light. This property of aequorin has allowed for its as a bioluminescent label in binding assays for a variety of analytes. Due to the excellent detection limits we demonstrated in assays for small peptides using a fusion protein between the peptide of interest and the photoprotein, our next goal was to expand the range of possible analytes for producing homogeneous populations of conjugates with the aequorin label to those that were nonpeptidic in nature. Recently, we prepared and characterized four aequorin mutants containing unique cysteine residues at various positions in the polypeptide chain. In the work reported here, the four aequorin mutants were each conjugated with a maleimide-activated methyl ester derivative of thyroxine, a hormone frequently determined to evaluate thyroid function. The thyroxine-aequorin mutant conjugates were characterized in terms of the bioluminescence activities and binding properties with an anti-thyroxine monoclonal antibody for possible future employment in either heterogeneous or homogeneous binding assays for thyroxine and/or other desired analytes.

UR - http://www.scopus.com/inward/record.url?scp=0034073753&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0034073753&partnerID=8YFLogxK

U2 - 10.1021/bc990081s

DO - 10.1021/bc990081s

M3 - Article

C2 - 10725089

AN - SCOPUS:0034073753

VL - 11

SP - 140

EP - 145

JO - Bioconjugate Chemistry

JF - Bioconjugate Chemistry

SN - 1043-1802

IS - 2

ER -