ABSTRACT: BACKGROUND:Due to a multitude of industrial applications of keratinolytic proteases, their demands are increasing. The present investigation studied the production and monitoring of the most possible multi-functional applications of YNDH thermoalkaline keratin-degrading enzyme. RESULTS:This work is considered the first that reported YNDH strain closely related to Laceyella sacchari strain; YNDH is a producer of protease/keratinase enzyme and able to degrade natural keratin such as feathers, wool, human hairs, and nails. Experimental design Plackett-Burman (PBD) was applied to evaluate culture conditions affecting the production of thermoalkaline protease/keratinase. Afterwards, Box-Behnken design (BBD) was applied to find out the optimum level of significant variables namely, NH4Cl, yeast extract, and NaNO3 with a predicted activity of 1324.7?U/ml. Accordingly, the following medium composition and parameters were calculated to be optimum (%w/v): NH4Cl, 0.08; feather, 1; yeast extract, 0.04; MgSO4.7H2O, 0.02; NaNO3, 0.016; KH2PO4, 0.01; K2HPO4, 0.01; pH, 8; inoculum size; 5%, cultivation temperature (Temp.) 45?°C and incubation time 48?h. The studied enzyme can degrade keratin-azure, remove proteinaceous materials, and is able to remove hairs from goat hides. These interesting characteristics make this enzyme a good candidate in many applications especially in detergent (Det.), in leather industries, and in pharmaceuticals particularly in nail treatment. CONCLUSION:The promising properties of the newly keratin-degrading protease enzyme from Laceyella sacchari strain YNDH would underpin its efficient exploitation in several industries to cope with the demands of worldwide enzyme markets.