?HIGH MOLECULAR WEIGHT ACID PHOSPHATASE FROM CHICKEN LIVER: ISOLATION, PuRIFICATION AND CHARACTERIZATION

Abstract

High molecular weight acid phosphatase from chicken liver was purified by salt fractionation, gel filtration on
Sephadex G-150 and by affinity chromatography on Sepharose4B-L-Tartramicamide column. The purified enzyme
moved as a single band. The molecular weight was estimated to be approximately 100 k.Oa. by gel filtration on
SephadexG-l 00andapproximately50k.Oa.by SOSpolyacrylamidegelelectrophoresisindicatingdimericprotein.K,
againstp-nitrophenyl phosphate and phenyl phosphate at pH 5.0 and ionic strength 0.15 were found to be 0.08 mM
and 0.12mM respectively, while the Vmax for these two substrates were 275 Umin' mg' and 250 Umin' mg! of
protein respectively. The enzyme was strongly inhibited by phosphate, tartrate, vanadate and molybdate. The ideal
substrates for the enzyme were4-ethyl phenyl phosphate,4-trifluoromethy-phenyl phosphate, a-naphthyl phosphate
and o-phosphotyrosine in addition to p-nitrophenyl phosphate and phenyl phosphate. Other substrates were also
hydrolysed but at slower rates.