Three-step purification of gp96 from human liver tumor tissues suitable for isolation of gp96-bound peptides

Song-Dong Meng; Jian Song; Zihe Rao; Po Tien; George F Gao

doi:10.1016/s0022-1759(02)00093-5

Three-step purification of gp96 from human liver tumor tissues suitable for isolation of gp96-bound peptides

J Immunol Methods. 2002 Jun 1;264(1-2):29-35. doi: 10.1016/s0022-1759(02)00093-5.

Authors

Song-Dong Meng¹, Jian Song, Zihe Rao, Po Tien, George F Gao

Affiliation

¹ Department of Molecular Virology, Institute of Microbiology, Chinese Academy of Sciences, Beijing, PR China.

PMID: 12191506
DOI: 10.1016/s0022-1759(02)00093-5

Abstract

Glycoprotein 96 (gp96), a member of heat shock protein (HSP) family, plays important roles in both innate immunity by itself and in T cell adaptive immunity in complex with antigenic-specific peptides. Using ammonium sulfate precipitation, ConA-Sepharose affinity chromatography and anion exchange chromatography, we have successfully purified gp96 from human liver tumor tissues. Subsequently, the gp96-associated peptides were successfully isolated from the gp96 preparation in quantities sufficient for micro-sequencing and identification of the peptides [Lancet 357 (2001) 528]. This three-step purification method might be used as a universal technique for the easy and reproducible isolation of antigenic heat shock protein gp96 from liver tissue or even other tissues of different sources. This will inevitably trigger the search for antigenic-specific peptides bound to gp96. Here, we report the method in detail.

Publication types

Research Support, Non-U.S. Gov't
Validation Study

MeSH terms

Ammonium Sulfate
Antigens, Neoplasm / isolation & purification*
Antigens, Neoplasm / metabolism*
Base Sequence
Carcinoma, Hepatocellular / chemistry*
Carcinoma, Hepatocellular / virology
Chemical Precipitation
Chromatography, Affinity / methods
Chromatography, Ion Exchange / methods
HSP70 Heat-Shock Proteins / isolation & purification*
HSP70 Heat-Shock Proteins / metabolism*
Hepatitis B Core Antigens / isolation & purification
Hepatitis B Core Antigens / metabolism
Hepatitis B virus / immunology
Humans
Liver Neoplasms / chemistry*
Liver Neoplasms / virology
Membrane Proteins / isolation & purification*
Membrane Proteins / metabolism*
Molecular Sequence Data
Neoplasm Proteins / isolation & purification
Neoplasm Proteins / metabolism
Peptides / isolation & purification*
Peptides / metabolism*
Protein Binding
Sequence Analysis, DNA
Sequence Analysis, Protein

Substances

Antigens, Neoplasm
HSP70 Heat-Shock Proteins
Hepatitis B Core Antigens
Membrane Proteins
Neoplasm Proteins
Peptides
glucose-regulated proteins
sarcoma glycoprotein gp96 rejection antigens
Ammonium Sulfate