Many proteins can easily attain a non-native fold and be of no use or even a
detriment to the host. The host cell has a myriad of molecules dedicated to assisting
nascent and existing proteins in folding properly and maintaining the native fold. Of
these molecular chaperones, the small Heat Shock Proteins (sHSP’s) are an important
group and worthy of study. The sHSP’s are a diverse group of proteins that have in
common an a-crystallin domain and generally display a chaperone activity. A sHSP
(HSP20) isolated from the cattle parasite Babesia bovis has similar activities, and limited
sequence homology to other a-crystallins. The gene encoding HSP20 was cloned into an
expression system where the gene product was induced and purified for study. It was
shown that HSP20 inhibits thermally induced aggregation of alcohol dehydrogenase at
equimolar ratios. HSP20 was also used to significantly reduce amyloid formation of the
b-Amyloid (1-40) Peptide in vitro at the sub-stoichiometric ratio of 1:10. A study of the
oligomeric forms of HSP20 using size exclusion chromatography and gel electrophoresis
revealed a broad range of multimers present in solution. The distribution of oligomers
was affected by altering the solution conditions and concentration of the protein. The
domains responsible for multimerization of HSP20 were mapped via sequence homology with known a-crystallins. These regions correspond to 12 carboxy-terminal
amino acids and 50 amino-terminal amino acids. Truncated versions of HSP20 lacking
these proposed oligomerization domains were created using PCR of the original gene
and cloning into an expression vector as before. Using size exclusion chromatography,
gel electrophoresis and analytical centrifugation, we show that the deleted domains alter
the multimeric population of the protein in solution. The carboxy-terminal domain has a
slight effect on multimerization while the amino-terminal deletion results in a drastic
reduction in any multimers above a dimer under the conditions tested. Despite this
drastic change in the multimerization of HSP20, there were no changes in the activities
observed when compared to the full-length form. From this we conclude that the regions
responsible for multimerization play little role in the observed activities of HSP20
