Expression and analysis of recombinant Amb a V and Amb t V allergens. Comparison with native proteins by immunological assays and NMR spectroscopy

T. Rafnar, B. Ghosh, W. J. Metzler, S. K. Huang, M. P. Perry, L. Mueller, D. G. Marsh

Research output: Contribution to journalArticle

18 Citations (Scopus)

Abstract

The Amb V allergens are small, highly disulfide-bonded ragweed pollen allergens that serve as useful models for understanding the molecular basis of the human immune response. We have produced recombinant Amb a V and Amb t V (from short and giant ragweed pollens, respectively) in Escherichia coli and have compared their structural and functional characteristics to those of the native proteins. Recombinant Amb t V was indistinguishable from native Amb t V as determined by NMR spectroscopy and antibody-binding studies. Whereas inhibition analysis showed that recombinant Amb a V possessed only ~50% of the antibody-binding activity of native Amb a V, the two proteins were similarly effective in stimulating Amb a V-specific T-cells. Our results demonstrate that even highly homologous proteins exhibit different abilities to fold into their native three-dimensional conformations and establish the potential and limits of expressing the recombinant Amb V allergens intracellularly in E. coli.

Original languageEnglish
Pages (from-to)21119-21123
Number of pages5
JournalJournal of Biological Chemistry
Volume267
Issue number29
Publication statusPublished - Jan 1 1992
Externally publishedYes

Fingerprint

Allergens
Nuclear magnetic resonance spectroscopy
Assays
Magnetic Resonance Spectroscopy
Escherichia coli
Proteins
Molecular Models
T-cells
Antibodies
Disulfides
Conformations
T-Lymphocytes
ragweed pollen

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Expression and analysis of recombinant Amb a V and Amb t V allergens. Comparison with native proteins by immunological assays and NMR spectroscopy. / Rafnar, T.; Ghosh, B.; Metzler, W. J.; Huang, S. K.; Perry, M. P.; Mueller, L.; Marsh, D. G.

In: Journal of Biological Chemistry, Vol. 267, No. 29, 01.01.1992, p. 21119-21123.

Research output: Contribution to journalArticle

Rafnar, T. ; Ghosh, B. ; Metzler, W. J. ; Huang, S. K. ; Perry, M. P. ; Mueller, L. ; Marsh, D. G. / Expression and analysis of recombinant Amb a V and Amb t V allergens. Comparison with native proteins by immunological assays and NMR spectroscopy. In: Journal of Biological Chemistry. 1992 ; Vol. 267, No. 29. pp. 21119-21123.
@article{5b3cf67afd2048f7bc77f95534d1ede9,
title = "Expression and analysis of recombinant Amb a V and Amb t V allergens. Comparison with native proteins by immunological assays and NMR spectroscopy",
abstract = "The Amb V allergens are small, highly disulfide-bonded ragweed pollen allergens that serve as useful models for understanding the molecular basis of the human immune response. We have produced recombinant Amb a V and Amb t V (from short and giant ragweed pollens, respectively) in Escherichia coli and have compared their structural and functional characteristics to those of the native proteins. Recombinant Amb t V was indistinguishable from native Amb t V as determined by NMR spectroscopy and antibody-binding studies. Whereas inhibition analysis showed that recombinant Amb a V possessed only ~50{\%} of the antibody-binding activity of native Amb a V, the two proteins were similarly effective in stimulating Amb a V-specific T-cells. Our results demonstrate that even highly homologous proteins exhibit different abilities to fold into their native three-dimensional conformations and establish the potential and limits of expressing the recombinant Amb V allergens intracellularly in E. coli.",
author = "T. Rafnar and B. Ghosh and Metzler, {W. J.} and Huang, {S. K.} and Perry, {M. P.} and L. Mueller and Marsh, {D. G.}",
year = "1992",
month = "1",
day = "1",
language = "English",
volume = "267",
pages = "21119--21123",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "29",

}

TY - JOUR

T1 - Expression and analysis of recombinant Amb a V and Amb t V allergens. Comparison with native proteins by immunological assays and NMR spectroscopy

AU - Rafnar, T.

AU - Ghosh, B.

AU - Metzler, W. J.

AU - Huang, S. K.

AU - Perry, M. P.

AU - Mueller, L.

AU - Marsh, D. G.

PY - 1992/1/1

Y1 - 1992/1/1

N2 - The Amb V allergens are small, highly disulfide-bonded ragweed pollen allergens that serve as useful models for understanding the molecular basis of the human immune response. We have produced recombinant Amb a V and Amb t V (from short and giant ragweed pollens, respectively) in Escherichia coli and have compared their structural and functional characteristics to those of the native proteins. Recombinant Amb t V was indistinguishable from native Amb t V as determined by NMR spectroscopy and antibody-binding studies. Whereas inhibition analysis showed that recombinant Amb a V possessed only ~50% of the antibody-binding activity of native Amb a V, the two proteins were similarly effective in stimulating Amb a V-specific T-cells. Our results demonstrate that even highly homologous proteins exhibit different abilities to fold into their native three-dimensional conformations and establish the potential and limits of expressing the recombinant Amb V allergens intracellularly in E. coli.

AB - The Amb V allergens are small, highly disulfide-bonded ragweed pollen allergens that serve as useful models for understanding the molecular basis of the human immune response. We have produced recombinant Amb a V and Amb t V (from short and giant ragweed pollens, respectively) in Escherichia coli and have compared their structural and functional characteristics to those of the native proteins. Recombinant Amb t V was indistinguishable from native Amb t V as determined by NMR spectroscopy and antibody-binding studies. Whereas inhibition analysis showed that recombinant Amb a V possessed only ~50% of the antibody-binding activity of native Amb a V, the two proteins were similarly effective in stimulating Amb a V-specific T-cells. Our results demonstrate that even highly homologous proteins exhibit different abilities to fold into their native three-dimensional conformations and establish the potential and limits of expressing the recombinant Amb V allergens intracellularly in E. coli.

UR - http://www.scopus.com/inward/record.url?scp=0026703078&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0026703078&partnerID=8YFLogxK

M3 - Article

C2 - 1400422

AN - SCOPUS:0026703078

VL - 267

SP - 21119

EP - 21123

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 29

ER -