Detergent enhances binding of a secreted HLA-A2 molecule to solid phase peptides

Lynda G. Tussey, Jeffrey A Frelinger

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

We have constructed a secreted analogue (sA2) of the human class I molecule HLA-A2. sA2 was affinity purified both in the presence and absence of detergent and the effects of detergent on the magnitude and specificity of A2 binding to solid phase peptides tested. sA2 purified in the presence of detergent and detergent-solubilized A2 are shown to function comparably in the binding of the synthetic peptide M.Y + 57-68, a known T-cell epitope derived from the influenza A matrix protein. The molecules binding to M.Y + 57-68 typically represent 8% to 10% of the added protein. In contrast, less than 1% of sA2 protein purified in the absence of detergent binds M.Y + 57-68. This reduced binding is not due to a change in the affinity of sA2 for M.Y + 57-68. Addition of detergent at various stages of the purification and iodination procedures indicates that the longer the sA2 molecules are exposed to detergent the better they bind. However, the concentration of detergent during the actual binding assay does not appear to be critical. We also find that while the sA2-detergent and the sA2-no detergent molecules differ in the extent to which they bind various peptides, they do not differ in their patterns of binding. We conclude that detergent probably does not influence the specificity of class I/peptide binding but does increase the number of sA2 molecules that can participate in the binding of peptide either by generating and stabilizing "empty" sA2 molecules or by stabilizing a structure that is more amenable to binding peptide.

Original languageEnglish (US)
Pages (from-to)183-193
Number of pages11
JournalHuman Immunology
Volume32
Issue number3
DOIs
StatePublished - 1991
Externally publishedYes

Fingerprint

HLA-A2 Antigen
Detergents
Peptides
varespladib methyl
Proteins
T-Lymphocyte Epitopes
Halogenation
Human Influenza

ASJC Scopus subject areas

  • Immunology
  • Immunology and Allergy

Cite this

Detergent enhances binding of a secreted HLA-A2 molecule to solid phase peptides. / Tussey, Lynda G.; Frelinger, Jeffrey A.

In: Human Immunology, Vol. 32, No. 3, 1991, p. 183-193.

Research output: Contribution to journalArticle

@article{596c95e4e2be4097aa25fca18de40fe8,
title = "Detergent enhances binding of a secreted HLA-A2 molecule to solid phase peptides",
abstract = "We have constructed a secreted analogue (sA2) of the human class I molecule HLA-A2. sA2 was affinity purified both in the presence and absence of detergent and the effects of detergent on the magnitude and specificity of A2 binding to solid phase peptides tested. sA2 purified in the presence of detergent and detergent-solubilized A2 are shown to function comparably in the binding of the synthetic peptide M.Y + 57-68, a known T-cell epitope derived from the influenza A matrix protein. The molecules binding to M.Y + 57-68 typically represent 8{\%} to 10{\%} of the added protein. In contrast, less than 1{\%} of sA2 protein purified in the absence of detergent binds M.Y + 57-68. This reduced binding is not due to a change in the affinity of sA2 for M.Y + 57-68. Addition of detergent at various stages of the purification and iodination procedures indicates that the longer the sA2 molecules are exposed to detergent the better they bind. However, the concentration of detergent during the actual binding assay does not appear to be critical. We also find that while the sA2-detergent and the sA2-no detergent molecules differ in the extent to which they bind various peptides, they do not differ in their patterns of binding. We conclude that detergent probably does not influence the specificity of class I/peptide binding but does increase the number of sA2 molecules that can participate in the binding of peptide either by generating and stabilizing {"}empty{"} sA2 molecules or by stabilizing a structure that is more amenable to binding peptide.",
author = "Tussey, {Lynda G.} and Frelinger, {Jeffrey A}",
year = "1991",
doi = "10.1016/0198-8859(91)90055-E",
language = "English (US)",
volume = "32",
pages = "183--193",
journal = "Human Immunology",
issn = "0198-8859",
publisher = "Elsevier Inc.",
number = "3",

}

TY - JOUR

T1 - Detergent enhances binding of a secreted HLA-A2 molecule to solid phase peptides

AU - Tussey, Lynda G.

AU - Frelinger, Jeffrey A

PY - 1991

Y1 - 1991

N2 - We have constructed a secreted analogue (sA2) of the human class I molecule HLA-A2. sA2 was affinity purified both in the presence and absence of detergent and the effects of detergent on the magnitude and specificity of A2 binding to solid phase peptides tested. sA2 purified in the presence of detergent and detergent-solubilized A2 are shown to function comparably in the binding of the synthetic peptide M.Y + 57-68, a known T-cell epitope derived from the influenza A matrix protein. The molecules binding to M.Y + 57-68 typically represent 8% to 10% of the added protein. In contrast, less than 1% of sA2 protein purified in the absence of detergent binds M.Y + 57-68. This reduced binding is not due to a change in the affinity of sA2 for M.Y + 57-68. Addition of detergent at various stages of the purification and iodination procedures indicates that the longer the sA2 molecules are exposed to detergent the better they bind. However, the concentration of detergent during the actual binding assay does not appear to be critical. We also find that while the sA2-detergent and the sA2-no detergent molecules differ in the extent to which they bind various peptides, they do not differ in their patterns of binding. We conclude that detergent probably does not influence the specificity of class I/peptide binding but does increase the number of sA2 molecules that can participate in the binding of peptide either by generating and stabilizing "empty" sA2 molecules or by stabilizing a structure that is more amenable to binding peptide.

AB - We have constructed a secreted analogue (sA2) of the human class I molecule HLA-A2. sA2 was affinity purified both in the presence and absence of detergent and the effects of detergent on the magnitude and specificity of A2 binding to solid phase peptides tested. sA2 purified in the presence of detergent and detergent-solubilized A2 are shown to function comparably in the binding of the synthetic peptide M.Y + 57-68, a known T-cell epitope derived from the influenza A matrix protein. The molecules binding to M.Y + 57-68 typically represent 8% to 10% of the added protein. In contrast, less than 1% of sA2 protein purified in the absence of detergent binds M.Y + 57-68. This reduced binding is not due to a change in the affinity of sA2 for M.Y + 57-68. Addition of detergent at various stages of the purification and iodination procedures indicates that the longer the sA2 molecules are exposed to detergent the better they bind. However, the concentration of detergent during the actual binding assay does not appear to be critical. We also find that while the sA2-detergent and the sA2-no detergent molecules differ in the extent to which they bind various peptides, they do not differ in their patterns of binding. We conclude that detergent probably does not influence the specificity of class I/peptide binding but does increase the number of sA2 molecules that can participate in the binding of peptide either by generating and stabilizing "empty" sA2 molecules or by stabilizing a structure that is more amenable to binding peptide.

UR - http://www.scopus.com/inward/record.url?scp=0026048365&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0026048365&partnerID=8YFLogxK

U2 - 10.1016/0198-8859(91)90055-E

DO - 10.1016/0198-8859(91)90055-E

M3 - Article

C2 - 1774199

AN - SCOPUS:0026048365

VL - 32

SP - 183

EP - 193

JO - Human Immunology

JF - Human Immunology

SN - 0198-8859

IS - 3

ER -