| The crustacean, Artemia franciscana, undergoes encystment and diapause, exhibiting a remarkable resistance to physiological stresses. Encysted Artemia embryos contain an abundant molecular chaperone termed artemin, thought to contribute to development and survival. To investigate the role of artemin during Artemia development and upon exposure to stress, the structural and functional characteristics of wild type and modified artemin were examined. Artemin mutants were generated by site-directed mutagenesis. Oligomer formation and chaperone activity were examined to study artemin function. Modified artemin had decreased chaperoning capability, with ArtC172A most dramatically affected. Oligomerization may be necessary but not sufficient for full chaperone activity, since all mutants formed large oligomers while exhibiting decreased chaperone capability. Computer modeling indicated that the four cysteines examined in this study are potentially located in a hydrophobic region, and that two of these are on the surface of the artemin oligomer. Interestingly, cysteine 172 was not only located in the hydrophobic region, but also associated with apparent pores in the protein shell of the oligomer, indicating this portion of the protein complex is part of the chaperone active site. This work has important implications for understanding the relationship between artemin structural organization and functional mechanisms during Artemia development and upon exposure to stress. |
