Abstract
The temperature sensitivity of crude solubilized 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, prepared from rat liver microsomes by different procedures, varies widely. When enzyme was solubilized by several techniques, HMG-CoA reductase activity in these extracts was rapidly and irreversibly inactivated at 4 degrees (Brown, M.S. Dana, S.E. Dietschy, J.M. and Siperstein, M.D. (1973) J. Biol. Chem 248, 4731-4738). In contrast, HMG-CoA reductase, solubilized from microsomes by a slow freeze-thaw method, was reversibility inactivated at 4 degrees over an interval of 2 h (Heller, R.A. and Gould, R.G. (1974) J. Biol. Chem 249, 5254-5260). In the present article irreversible inactivation at 4 degrees of crude solubilized HMG-CoA reductase, prepared from liver microsomes by a slow freeze-thaw technique, was investigated. In the absence of preincubation at 37 degrees, HMG-CoA reductase activity in the crude soluble extract had a half-life at 4 degrees of 4 h. Enzyme activity was more rapidly inactivated (t1/2=4 h) in extracts from younger (150 to 250 g) rats than in extracts from older (500 g) rats (t1/2=37 h). In contrast, partially purified HMG-CoA reductase was far more stable at 4 degrees (t1/2=312 h). However, when partially purified enzyme was treated with crude soluble extract, the enzyme was inactivated much more rapidly (t1/2=26 h). It is concluded that rapid irreversible inactivation of HMG-CoA reductase at 4 degrees is not an intrinsic property of this enzyme, but instead, this inactivation is caused by a factor or factors present in the crude soluble extract. While HMG-CoA reductase activity was rapidly inactivated in crude extracts from animals killed at the midpoint of the dark cycle, enzyme activity was inactivated much more slowly in extracts obtained from animals killed at the midpoint of the light cycle. These findings suggest that the concentration of the inactivating factor may vary, depending upon the physiological state of the animal. The nature of the inactivating factor is not known at the present time; however, it may be a protein since it is nondialyzable and destroyed by heat.
Highlights
It is concluded that rapid irreversible inactivation of hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase at 4” is not an intrinsic property of this enzyme, but instead, this inactivation is caused by a factor or factors present in the crude soluble extract
Reductase diluted with soluble extract was D&fold more rapidly inactivated than partially purified enzyme diluted with Buffer A. These results show that rapid irreversible inactivation of HMG-CoA reductase at 4” is not an intrinsic property of this enzyme, but instead, this rapid inactivation is caused by a factor or factors present in the crude soluble extract which are either not present or present at greatly reduced concentration in the partially purified enzyme
The temperature sensitivity of crude solubilized preparations of HMG-CoA reductase prepared by different procedures varies widely [11]
Summary
When enzyme was solubilized by several techniques, HMG-CoA reductase activity in these extracts was rapidly and irreversibly inactivated at 4” In the present article irreversible inactivation at 4” of crude solubilized HMG-CoA reductase, prepared from liver microsomes by a slow freeze-thaw technique, was investigated. When partially purified enzyme was treated with crude soluble extract, the enzyme was inactivated much more rapidly (t,,2 = 26 h). While HMG-CoA reductase activity was rapidly inactivated in crude extracts from animals killed at the midpoint of the dark cycle, enzyme activity was inactivated much more slowly in extracts obtained from animals killed at the midpoint of the light cycle These findings suggest that the concentration of the inactivating factor may vary, depending upon the physiological state of the animal. HMG-CoA reductase solubilized by the slow freeze-thaw method of Heller and
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
