Improved procedure for the purification of hepatic lipase from rat liver homogenate.
A procedure is described for the purification of hepatic lipase (HL)4 from rat liver homogenate which results in a high yield (41%) of electrophoretically homogeneous enzyme. The method is based on that of Twu et al. (Biochim. Biophys. Acta 1984: 792, 330), but it is more efficient with respect to yield (about 4-fold) and purity (1.6-fold). It includes the preparation of a high-speed supernatant, chromatography in series on octyl-, heparin- and concanavalin A-Sepharose, and finally gel filtration. On SDS-PAGE analysis, the purified enzyme exhibited an apparent molecular mass of 63.6 +/- 3.2 kDa. Heterogeneity was observed, when purified HL was subjected to isoelectric focussing. The enzyme displayed a specific catalytic activity of 23,000 U* (mumol fatty acid released per h at 37 degrees C) per mg protein, when assayed with trioleoyl glycerol suspensions in arabic gum. A highly specific antiserum against rat liver HL, capable of inhibiting 817 mU* HL per microliter antiserum, was raised in rabbits.
Kresse
H
H
Max-Planck-Institut für Experimentelle Medizin, Göttingen, Germany.
Löffler
B M
BM
Kunze
H
H
D000818:Animals; D002846:Chromatography, Affinity; D002847:Chromatography, Agarose; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D005227:Fatty Acids / Q000032:analysis / Q000378:metabolism; D007525:Isoelectric Focusing; D008049:Lipase / Q000032:analysis / Q000302:isolation & purification* / Q000378:metabolism; D008099:Liver / Q000201:enzymology*; D008297:Male; D008970:Molecular Weight; D051381:Rats; D011919:Rats, Inbred Strains
D005227:Fatty Acids; D008049:Lipase