Proteolytic specificity of chymosin on bovine αs1-casein

Abstract

SummaryIn dilute buffers ⋟ pH 5·8, chymosin hydrolysed bovine αs1-casein to αs1-I, αs1-II and αs1-III/αs1-IV in a sequential manner while at pH 4·6 αs1-casein was hydrolysed to αs1-I which was then hydrolysed to αs1-V. In the presence of 5 % (w/v) NaCl at pH 5·2, αs1-casein was hydrolysed to αs1-I which was then hydrolysed to αs1-VII and αs1-VIII. αs1-I, αs1-II and αs1-III/αs1-IV were isolated by chromatography on cellulose phosphate followed by preparative slab-gel electrophoresis; αs1-V was isolated by repeated preparative slab-gel electrophoresis and αs1-VII by gel filtration on Sephadex G-150 followed by preparative slab-gel electrophoresis. The mol. wts of the peptides, estimated by gel filtration on Sephadex G-100, were 21000, 17600, 15600, 19900 and 14600 for αs1-I, αs1-II, αs1-III/αs1-IV and αs1-V and αs1-VII respectively. Characterization of the peptides by amino acid, phosphorus and terminal residue analysis showed that they probably consisted of segments of the αs1-casein chain as follows: αs1-I: residues 24/25–199; αs1-II: residues 24/25–169; αs1-III/αs1-IV: residues 24/25–149–150; αs1-V: residues 29/33–199; αs1-VII: residues 56–179. Peptide bonds close to phosphate residues on the αs1-casein chain were not hydrolysed by chymosin at high pH values (⋟ 5·8) when the phosphate groups were charged, but became available for hydrolysis when the reaction pH was reduced. Proteolytic specificity was also modified by NaCl.