Subtilisins of Bacillus spp. hydrolyze keratin and allow growth on feathers

Keratinase is a serine protease produced by Bacillus licheniformis PWD-1 th at effectively degrades keratin and confers the ability to grow on feathers to a protease-deficient B. subtilis strain. Studies presented herein demon strate that B. licheniformis Carlsberg strain NCIMB 6816, which produces th e well-characterized serine protease subtilisin Carlsberg, also degrades an d grows on feathers. The PWD-1 and Carlsberg strains showed a similar time- course of enzyme production, and the purified serine proteases have similar enzymatic properties on insoluble azokeratin and soluble FITC-casein. Kine tic analysis of both enzymes demonstrated that they have high specificity f or aromatic and hydrophobic amino acids in the P1 substrate position, altho ugh keratinase discriminates more than subtilisin Carlsberg against charged residues at this site. Nucleotide sequence analysis of the serine protease genes from B. licheniformis strains PWD-1, Carlsberg NCIMB 6816, ATCC 1275 9, and NCIMB 10689 showed that the kerA-encoded protease of PWD-1 differs f rom the others only by having V222, rather than A222, near the active site serine S220. Further, high-level expression of subE-encoded subtilisin from B. subtilis (78% similar to subtilisin Carlsberg) also confers growth on f eathers on a protease-deficient B. subtilis strain. While strain PWD-1 and the kerA protease efficiently degrade keratin, keratin hydrolysis and growt h on feathers is a property that can be conferred by appropriate expression of the major subtilisins, including the industrially produced enzymes.