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Characterization of the 16 kDalton heat shock proteins of Caenorhabditis elegans Hockertz, Michael Karl


The 16 kDa heat shock proteins of C. elegans have been partially purified and characterized using antipeptide antibodies and standard chromatographic techniques. Based upon the sequences of the previously isolated hsp16 genes, peptides were synthesized against which polyclonal antibodies were produced in rabbits. The antibodies were used to detect fractionated hspl6 by the western blot technique and to purify hsp16 by affinity chromatography. The specificity of the anti-peptide antibodies indicated that the four hsp16 genes may encoded both 16 kDa and the 18 kDa polypeptides. The latter were previously thought to be derived from distinct genes. Two-dimensional gel electrophoresis showed that the 16 kDa hsps exist in multiple isoforms, probably due to their posttranslational modification. Hsp16 was purified by affinity chromatography with immobilized antibody to a C-terminal 36 residue peptide of hsp16-2. The hspl6s did not bind to the antibody in a low salt buffer, but did so in the presence 4M urea. Hsp16 was fractionated by hydroxylapatite and gel exclusion chromatography and shown to exist as a large reasonably uniform complex of 460±55 kDa.

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