title: The role of the sequence extensions in beta-crystallin assembly.
abstract: 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 beta-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.
authors.affiliation: Department of Molecular Biology, University of Nijmegen, The Netherlands.
authors.forename: R C
authors.initials: RC
authors.lastname: Kroone
authors.forename: G S
authors.initials: GS
authors.lastname: Elliott
authors.forename: A
authors.initials: A
authors.lastname: Ferszt
authors.forename: C
authors.initials: C
authors.lastname: Slingsby
authors.forename: N H
authors.initials: NH
authors.lastname: Lubsen
authors.forename: J G
authors.initials: JG
authors.lastname: Schoenmakers
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002942:Circular Dichroism; D003459:Crystallins / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D017510:Protein Folding; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism
chemical_list: D003459:Crystallins; D011994:Recombinant Proteins