Heterologous expression and characterisation of a keratinase produced by Chryseobacterium carnipullorum

Chryseobacterium carnipullorum 9_R23581^T, isolated from raw chicken meat, was evaluated for its potential to degrade keratin found in feathers. The focus of this study was to heterologously express and characterise a keratinolytic enzyme produced by C. carnipullorum. Chryseobacterium carnipullorum secretes proteolytic enzymes that have feather degrading capabilities during its exponential growth phase. This study concluded that the most likely main component of the keratinolytic enzymes of C. carnipullorum was peptidase M64, a serine-endopeptidase with a molecular weight in crude form of 49.46 kDa. Primers were designed on the selected gene of interest, which was amplified from the genome of C. carnipullorum (accession number NZ-FRCD01000002.1). The gene coding for peptidase M64 was further cloned, propagated and expressed in E. coli BL21 [DE3] cells. Purification was by Immobilised Metal Affinity Chromatography (IMAC). The molecular weight of the keratinase was about 50 kDa after purification while its optimum temperature and pH were 50 °C and 8.5, respectively. The activity of this keratinase was inhibited by phenylmethylsulfonyl fluoride (PMSF) and it was enhanced by the presence of divalent metal ions such as Mg²⁺ and Ca²⁺. Enzyme activity was further assayed by application to chicken feathers and observed degradation was an indication of keratinolytic potential.