Partial purification and characterization of hormonesensitive lipase from rat adipose tissue

Abstract

1. 1. A high yield of triglyceride lipase was recovered reproducibly in the clear 105000 × g supernatant (S 105 fraction) when rat epididymal fat pads were homogenized in 0.25 M sucrose containing 10 −3 M EDTA. 2. 2. Previous exposure of intact fat pads to epinephrine (5 · 10 −5 M) caused an increment in the triglyceride lipase activity recovered in the S 105 fraction. 3. 3. This hormone-sensitive lipase activity was not inhibited by 1 M NaCl or protamine sulfate (800 μg/ml). NaF (10 −2 M) inhibited the activity by 80%. 4. 4. A part of the lipase activity floated during preparative ultracentrifugation at medium densities greater than 1.125. Previous treatment of the intact fat pads with epinephrine increased the activity in this floating ( d < 1.125) fraction. 5. 5. Lipase activity in the S 105 fraction was rapidly inactivated during incubation at 37° and pH 7.4. This deactivation could be blocked by 10 −3 M EDTA. 6. 6. After 35-fold purification of lipase, obtained by precipitation at pH 5.2 and preparative ultracentrifugation at d = 1.125, the enzyme was stable even in the absence of EDTA. 7. 7. A high level of lipoprotein lipase activity was demonstrable in the S 105 fraction when fat pads from fed rats were used. Fasting of the animals or preincubation of fat pads in glucose-free medium lowered this activity without changing the activity of the hormone-sensitive triglyceride lipase.