Fish otoliths composed of calcium carbonate and an organic matrix play a primary role in gravity sensing and the perception of sound. Starmaker (Stm) was the first protein found to be capable of influencing the process of biomineralization of otoliths. Stm dictates the shape, size, and selection of calcium carbonate polymorphs in a concentration-dependent manner. To facilitate exploration of the molecular basis of Stm function, we have developed and optimized a protocol for efficient expression and purification of the homogeneous nontagged Stm. The homogeneous nontagged Stm corresponds to its functional form, which is devoid of a signal peptide. A comprehensive biochemical and biophysical analysis of recombinant Stm, along with in silico examinations, indicate for the first time that Stm exhibits the properties of intrinsically disordered proteins. The functional significance of Stm having intrinsically disordered protein properties and its possible role in controlling the formation of otoliths is discussed.

The Cdc25p and Sdc25p proteins were the first members of the family of guanine nucleotide exchange factors to be identified. These proteins promote the formation of active Ras-GTP complex from inactive Ras-GDP complex by exchange of GDP for GTP. Therefore Cdc25p which is the main positive regulator of Ras, regulates through Ras the activity of adenylate cyclase in Saccharomyces cerevisiae. The amino-terminal part of Cdc25p has a sequence similar to the cyclin destruction box (CDB) of mitotic cyclins. This sequence has been reported to be required for ubiquitin-dependent proteolysis. In this study we show that Cdc25p is an unstable polypeptide with a half-life of 15-20 min. Its instability depends upon the presence of the CDB which can also confer instability to other proteins. Degradation of Cdc25p and CDB containing beta-galactosidase was found to be independent of various cell cycle arrest points. The fast degradation of Cdc25p opens the possibility that Ras and the cAMP cascade in yeast are directly modulated by the cellular content of the guanine nucleotide exchange factor rather than variation in activity or localization control.

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