@article{chen_rojanatavorn_clark_shih_2005, title={Characterization and enzymatic degradation of Sup35NM, a yeast prion-like protein}, volume={14}, ISSN={["1469-896X"]}, DOI={10.1110/ps.041234405}, abstractNote={AbstractTransmissible spongiform encephalopathies (TSEs) are believed to be caused by an unconventional infectious agent, the prion protein. The pathogenic and infectious form of prion protein, PrPSc, is able to aggregate and form amyloid fibrils, very stable and resistant to most disinfecting processes and common proteases. Under specific conditions, PrPSc in bovine spongiform encephalopathy (BSE) brain tissue was found degradable by a bacterial keratinase and some other proteases. Since this disease‐causing prion is infectious and dangerous to work with, a model or surrogate protein that is safe is needed for the in vitro degradation study. Here a nonpathogenic yeast prion‐like protein, Sup35NM, cloned and overexpressed in E. coli, was purified and characterized for this purpose. Aggregation and deaggregation of Sup35NM were examined by electron microscopy, gel electrophoresis, Congo red binding, fluorescence, and Western blotting. The degradation of Sup35NM aggregates by keratinase and proteinase K under various conditions was studied and compared. These results will be of value in understanding the mechanism and optimization of the degradation process.}, number={9}, journal={PROTEIN SCIENCE}, author={Chen, CY and Rojanatavorn, K and Clark, AC and Shih, JCH}, year={2005}, month={Sep}, pages={2228–2235} }
 @article{wang_rojanatavorn_shih_2004, title={Increased production of Bacillus keratinase by chromosomal integration of multiple copies of the kerA gene}, volume={87}, ISSN={["0006-3592"]}, DOI={10.1002/bit.20145}, abstractNote={AbstractTo increase the production of keratinase, stable strains of Bacillus licheniformis carrying multiple keratinase gene copies in the chromosome were developed. Integrative vectors carrying kerA with or without P43‐promoter were constructed and subcloned into B. licheniformis T399D and Bacillus subtilis DB104. In T399D, multiple copies of kerA integration into the chromosome were identified and determined by Southern blot. The optimal integration of kerA was found in the range of 3–5 copies. Higher integration of gene copies (>5) caused reduced processing and secretion of the extracellular keratinase. In DB104, kerA was cloned in the plasmid, not integrated into the chromosome. The strong constitutive promoter P43 not only increased the keratinase production in plasmid‐based expression in DB104 but also improved the enzyme yield of the integrants of T399D. New strains were able to enhance cell growth and enzyme yield at higher concentrations of medium substrate. When they were grown in either soy or feather medium, the keratinase activity was stable and improved by about 4–6 times. © 2004 Wiley Periodicals, Inc.}, number={4}, journal={BIOTECHNOLOGY AND BIOENGINEERING}, author={Wang, JJ and Rojanatavorn, K and Shih, JCH}, year={2004}, month={Aug}, pages={459–464} }