Abstract
We describe here the purification and characterization of a form of acid lipase from human liver (designated ALII), which differed from the more abundant Mr-29000 form (ALI). ALII was solubilized from frozen human liver with Triton X-100 and purified 8500-fold by chromatography over concanavalin A-sepharose, CM-cellulose and finally h.p.l.c. over a Mono S column. ALII migrated as a single band on polyacrylamide-gel electrophoresis in both the presence and the absence of SDS. The Mr of ALII was estimated to be 58,500 by SDS/polyacrylamide-gel electrophoresis. Gel filtration on Sephacryl S-200 gave an apparent Mr of 69,000. 4-Methylumbelliferyl (4MU) palmitate, cholesterol oleate and triolein were substrates for ALII, with apparent Vmax values of 5000, 1100 and 2500 nmol/min per mg respectively and Km values of 1.0, 1.5 and 1.8 mM respectively. Cholesterol oleate and triolein were hydrolysed optimally by ALII at pH 4.5, whereas 4MU palmitate was hydrolysed optimally at pH 5.3. Antisera were raised against ALI and ALII and, on immunoblot analysis, no antigenic similarity was observed between ALI and ALII. Cellulose acetate electrophoresis followed by reaction with 4MU palmitate revealed two forms of lipase, corresponding to ALI and ALII. The two enzymes were also separated by hydrophobic chromatography. The activity of ALII was stimulated by several proteins and was partially inhibited by millimolar concentrations of NaCl, CaCl2 and MgSO4.