The role of the sequence extensions in beta-crystallin assembly
Kroone, R.C.; Elliott, G.S.; Ferszt, A.; Slingsby, C.; Lubsen, N.H.; Schoenmakers, J.G.
Protein Engineering 7(11): 1395-1399
1994
ISSN/ISBN: 0269-2139 PMID: 7700872 Document Number: 438259
The modular construction of the eye lens beta-gamma-crystallins makes them good candidates for protein engineering to ascertain the rules of assembly of oligomers. X-ray studies have shown that although the polypeptide chains of beta-B2-crystallin and gamma-crystallins fold to form similar N- and C-terminal domains, the conformation of the connecting peptides are such that the gamma-crystallins are monomers and the beta-crystallin is a dimer. Unlike gamma-crystallins, the numerous gamma-crystallins have extensions of variable sequence from the globular domains. We have tested the effect of removing the N- and C-terminal extensions from rat beta-B2-crystallin using a bacterial expression system. Abundant proteins were produced in Escherichia coli using the pET or pQE vectors. Full-length and truncated proteins were purified and checked for refolding using circular dichroism. Sizing of the truncated proteins using gel filtration chromatography showed that the absence of either the N- or C-terminal extension does not affect dimerization of beta-B2-crystallin.