A 43-fold increase in keratinase production by Kocuria rosea was achieved in batch fermentation using response surface methodology. Factorial designs were used to select the components of a culture medium that showed a significant effect on keratinase production. An orthogonal–central composite experimental design was performed, with only two (feathers and magnesium) from nine initial compounds being further analyzed by response surface methodology. An optimum keratinase production of 14 886.9 U/mg was obtained with the following medium composition (per litre): NH₄Cl, 0.3 g; NaCl, 0.3 g; K₂HPO₄, 3.2 g; KH₂PO₄, 4.0 g; MgSO₄·6H₂O, 0.5 g; yeast extract, 0.1 g; and finely milled feathers, 30 g. The medium was shaken at 400 r/min with an incubation period of 14 h at 40 °C.Key words: feathers, keratinases, Kocuria rosea, medium optimization, response surface methodology.