Structure of 85 kDa subunit of human phosphatidylinositol 3-kinase analyzed by using monoclonal antibodies

Autor: Michiyuki Matsuda, Satoshi Nagata, Kazuo Nagashima, Yasuhisa Fukui, Shinya Tanaka, Takeshi Kurata, Yasuhiko Shizawa
Rok vydání: 1993
Předmět:
Monoclonal antibody
Cancer Research
medicine.drug_class
Protein subunit
Immunoblotting
Molecular Sequence Data
Alpha (ethology)
macromolecular substances
Cross Reactions
Signal transduction
Peptide Mapping
Article
chemistry.chemical_compound
Epitopes
Mice
Phosphatidylinositol 3-Kinases
Structure-Activity Relationship
Chlorocebus aethiops
medicine
Animals
Humans
Tissue Distribution
Phosphatidylinositol
Cloning
Molecular
Mice
Inbred BALB C
Hybridomas
biology
Base Sequence
Phosphotransferases
Antibodies
Monoclonal
Molecular biology
Immunohistochemistry
Precipitin Tests
Recombinant Proteins
Epitope mapping
Phosphatidylinositol 3‐kinase
Oncology
chemistry
Biochemistry
Polyclonal antibodies
biology.protein
Antibody
hormones
hormone substitutes
and hormone antagonists
Polymorphism
Restriction Fragment Length
Zdroj: Japanese Journal of Cancer Research : Gann
ISSN: 0910-5050
Popis: An 85 kDa subunit (p85 alpha) of phosphatidylinositol 3-kinase (PI-3K) has one SH3 and two SH2 regions [SH2(N) and SH2(C)], which direct protein-protein interaction. We have established eighteen hybridomas producing monoclonal antibodies against p85 alpha to study the structure-function relationship of this protein. Epitope mapping using a series of deletion mutants expressed in E. coli showed that the monoclonal antibodies bound to at least 5 distinct epitope regions, which were well dispersed on p85 alpha except for its carboxyl-terminus. Monoclonal antibodies against amino-terminal regions and polyclonal antibodies against carboxyl-terminal regions immunoprecipitated p85 alpha expressed in human cells and in E. coli. On the other hand, monoclonal antibodies against the central part of p85 alpha failed to immunoprecipitate p85 alpha efficiently; however, they could immunoprecipitate p85 alpha mutants with deletion of either the amino- or the carboxyl-terminal region. Similar results were obtained by immunocytochemistry using confocal microscopy. These results suggested that steric hindrance prevents binding of monoclonal antibodies to the central part of p85 alpha where SH2(N) is located. The SH2(N) may have a distinct function from SH2(C), which is located at the carboxyl-terminal region and has been shown to mediate the binding of PI-3K to activated growth factor receptors.
Databáze: OpenAIRE
Externí odkaz: