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Production of recombinant soluble human integrin alpha4beta1.

FEBS letters (2000-04-18)
K Clark, P Newham, L Burrows, J A Askari, M J Humphries
ABSTRACT

Integrin alpha4beta1 is a major leukocyte adhesion receptor that is a key target for the development of anti-inflammatory therapeutics. With the dual long-term goals of developing a reagent for use in high-throughput inhibitor screening assays and for crystallisation trials and subsequent structure determination, we have generated a recombinant soluble alpha4beta1 receptor. Both subunits were truncated prior to the transmembrane domains by site-directed mutagenesis and expressed using baculovirus infection of insect cells. The molecular weights of the recombinant subunits were as expected for post-translationally unmodified protein. In addition, as observed for the native subunit, a proportion of the alpha4 subunit was proteolytically processed into two fragments. ELISA and solid phase ligand-binding assays were performed to investigate the folding and functionality of the soluble integrin. The data suggest that the receptor was correctly folded and that it bound recombinant ligands with similar kinetics to the native molecule.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Mimetic Green 1 PIKSI Column, suspension
Sigma-Aldrich
Mimetic Blue SA PIKSI Column, suspension
Sigma-Aldrich
Mimetic Blue 1 ALPHA Column, suspension
Sigma-Aldrich
Mimetic Blue 2 ALPHA Column, suspension
Sigma-Aldrich
Mimetic Blue AP PIKSI column, suspension
Sigma-Aldrich
Mimetic Red 2 PIKSI column, suspension
Sigma-Aldrich
Mimetic Red 3 ALPHA Column, suspension
Sigma-Aldrich
Mimetic Orange 1 PIKSI Column, suspension
Sigma-Aldrich
Mimetic Orange 3 ALPHA Column, suspension
Sigma-Aldrich
Mimetic Yellow 2 PIKSI Column, suspension
Sigma-Aldrich
Mimetic Blue 2 PIKSI Column, suspension
Sigma-Aldrich
Mimetic Red 2 ALPHA Column, suspension
Sigma-Aldrich
Mimetic Orange 3 PIKSI Column, suspension
Sigma-Aldrich
Mimetic Orange 1 ALPHA Column, suspension
Sigma-Aldrich
Mimetic Yellow 1 PIKSI Column, suspension
Sigma-Aldrich
Mimetic Green 1 ALPHA Column, suspension
Sigma-Aldrich
Mimetic Yellow 2 ALPHA Column, suspension