Graduate Thesis Or Dissertation
 

The effects of cyclopropenoid fatty acids on structural components of microsomal membranes

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  • Studies were conducted to determine the effects of cyclopropenoid fatty acids (CPFA) on the microsomal membrane of livers of rainbow trout (Salmo gairdneri). Slab and tube gel electrophoresis of microsomes from trout fed a CPFA diet (CPFA-microsomes) for varying time periods showed a decrease in the number of protein bands resolved in the high molecular weight region. This disappearance of high molecular weight proteins was not due to increased proteolysis in the CPFA-microsomes. Antibodies against whole microsomal protein from livers of trout fed 300 ppm CPFA were produced in rabbits. Microsomal proteins were first separated by polyacrylamide gel electrophoresis (PAGE), transferred to nitrocellulose sheets (NC) and analyzed by the peroxidase-antiperoxidase (PAP) immunochemical staining procedure. Immunoabsorption of antisera directed against CPFA-microsomes by control-microsomes did not reveal any new proteins induced by the CPFA diets. However, the intensity of PAP staining was much greater in CPFA microsomes after immunoabsorption. Hydrolysis of phospholipids in the microsomal membrane by phospholipase A₂ failed to reveal any differences between control and CPFA fed trout. Proteolysis of microsomal membrane proteins had similar effects on NADPH cytochrome reductase and cytochrome P-450 activity on fish fed the different diets. PAGE analysis of these digests did show some differences in digested proteins between the control and CPFA group. These results may reflect a possible change in orientation of microsomal membrane proteins brought about by CPFA in the diet. Additional evidence for altered orientation of proteins was found with PAGE analysis of trypsin-digested microsomes. Moreover incubation of trypsin-digested microsomes with antisera and stained with PAP showed that proteolytic attack was different between control and CPFA microsomes. A final study with incubation of transferred proteins from control and CPFA-microsomes with antisera directed against purified cytochrome P-450 (P-450) and cytochrome P-448 (P-448) showed that CPFA had an effect on the concentration of P-448 but not P-450.
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file details MORRISSEYMICHAEL1982.pdf 2017-08-19 公开 下载