The acid phosphatases of bovine leucocytes, plasma and the milk of healthy and mastitic cows

Abstract

Summary: Some of the acid phosphatase isozymes of bovine leucocytes and plasma have been separated and partly characterized. About 80% of the phosphatase activity of leucocytes at pH 4·9 was particle-bound and about 8% was extractable with Amberlite CG-50 ion exchange resin. This extractable enzyme existed as a single electrophoretic component with a mol. wt of about 42000 and with optimum activity at pH 5·8. K_mforp-nitrophenyl phosphate was 1·6 mM at pH 5·8 and 0·4 mM at pH 4·9. At pH 5·8 orthophosphate (K₁= 1·5 mM) and pyrophosphate (K_i= 4·1 mM) were competitive inhibitors. The enzyme was also strongly inhibited by F⁻, Al³⁺, IO₄⁻and S₂0₃²⁻. The enzyme which was not extractable with Amberlite was very heterogeneous with respect to molecular weight. At the pH optimum (4·9), K_mforp-nitrophenyl phosphate was 0·4 mM and orthophosphate (K₁= 2·3 mM) and pyrophosphate (K₁= 2·1 mM) were competitive inhibitors. Other inhibitors included F⁻, Al³⁺, Hg²⁺, IO₄⁻and tartrate. The enzyme extracted from plasma by Amberlite CG-50 treatment had properties similar to that extracted from leucocytes. Normal bovine milk contained a single acid phosphatase, but milk from cows with mastitis showed 3 electrophoretic isozyme bands, one being the same as in normal milk; the 2 additional bands were of leucocyte origin.