scholarly journals Hepatic mitochondrial inner membrane properties and carnitine palmitoyltransferase A and B. Effect of diabetes and starvation

1985 ◽  
Vol 232 (2) ◽  
pp. 445-450 ◽  
Author(s):  
L J Brady ◽  
L J Silverstein ◽  
C L Hoppel ◽  
P S Brady
Keyword(s):  
Membrane Fluidity ◽  
Diabetic Rats ◽  
Carnitine Palmitoyltransferase ◽  
Membrane Properties ◽  
Inner Membrane ◽  
Fed State ◽  
Treatment Groups ◽  
Mitochondrial Inner Membrane ◽  
Malonyl Coa ◽  
Carnitine Palmitoyltransferase Activity

Intact mitochondria and inverted submitochondrial vesicles were prepared from the liver of fed, starved (48 h) and streptozotocin-diabetic rats in order to characterize carnitine palmitoyltransferase kinetics and malonyl-CoA sensitivity in situ. In intact mitochondria, both starved and diabetic rats exhibited increased Vmax., increased Km for palmitoyl-CoA, and decreased sensitivity to malonyl-CoA inhibition. Inverted submitochondrial vesicles also showed increased Vmax. with starvation and diabetes, with no change in Km for either palmitoyl-CoA or carnitine. Inverted vesicles were uniformly less sensitive to malonyl-CoA regardless of treatment, and diabetes resulted in a further decrease in sensitivity. In part, differences in the response of carnitine palmitoyltransferase to starvation and diabetes may reside in differences in the membrane environment, as observed with Arrhenius plots, and the relation of enzyme activity and membrane fluidity. In all cases, whether rats were fed, starved or diabetic, and whether intact or inverted vesicles were examined, increasing membrane fluidity was associated with increasing activity. Malonyl-CoA was found to produce a decrease in intact mitochondrial membrane fluidity in the fed state, particularly at pH 7.0 or less. No effect was observed in intact mitochondria from starved or diabetic rats, or in inverted vesicles from any of the treatment groups. Through its effect on membrane fluidity, malonyl-CoA could regulate carnitine palmitoyltransferase activity on both surfaces of the inner membrane through an interaction with only the outer surface.

scholarly journals Hepatic mitochondrial inner-membrane properties, β-oxidation and carnitine palmitoyltransferases A and B. Effects of genetic obesity and starvation

1986 ◽  
Vol 233 (2) ◽  
pp. 427-433 ◽  
Author(s):  
L J Brady ◽  
C L Hoppel ◽  
P S Brady
Keyword(s):  
Membrane Fluidity ◽  
Carnitine Palmitoyltransferase ◽  
Membrane Properties ◽  
Zucker Rats ◽  
Beta Oxidation ◽  
Inner Membrane ◽  
Membrane Polarization ◽  
Mitochondrial Inner Membrane ◽  
Malonyl Coa ◽  
Obese Rats

Hepatic mitochondrial carnitine palmitoyltransferase (CPT) properties, beta-oxidation of palmitoyl-CoA and membrane polarization were measured in lean and obese Zucker rats. The Vmax. of the ‘outer’ carnitine palmitoyltransferase (‘CPT-A‘) increased with starvation, with no change in the Km for either carnitine or palmitoyl-CoA. The Ki for malonyl-CoA increased with starvation in lean rats, but not in obese rats. The Vmax. of the ‘inner’ enzyme (‘CPT-B‘), as measured by using inverted submitochondrial vesicles, increased with starvation in obese rats only, with no change in the Km for either carnitine or palmitoyl-CoA. The Ki for malonyl-CoA was 2-5-fold higher in inverted vesicles than in intact mitochondria, and showed no alteration with starvation. The activities of both enzymes correlated positively with each other and with beta-oxidation, and inversely with membrane polarization. Malonyl-CoA had little effect on gross membrane fluidity in the Zucker rat, as reflected by diphenylhexatriene fluorescence polarization. The results indicate that both enzymes are related and respond similarly to alterations in membrane fluidity. Membrane fluidity may provide a mechanism for co-ordinated control of CPT activity on both sides of the mitochondrial inner membrane.

scholarly journals Altered release of carnitine palmitoyltransferase activity by digitonin from liver mitochondria of rats in different physiological states

1985 ◽  
Vol 230 (2) ◽  
pp. 389-394 ◽  
Author(s):  
V A Zammit ◽  
C G Corstorphine
Keyword(s):  
Liver Mitochondria ◽  
Diabetic Rats ◽  
Carnitine Palmitoyltransferase ◽  
Rat Liver Mitochondria ◽  
Intermembrane Space ◽  
Marker Enzyme ◽  
Inner Membrane ◽  
Mitochondrial Inner Membrane ◽  
Cpt I ◽  
Carnitine Palmitoyltransferase Activity

The release of carnitine palmitoyltransferase (CPT) activity from rat liver mitochondria by increasing concentrations of digitonin was studied for mitochondrial preparations from fed, 48 h-starved and diabetic animals. A bimodal release of activity was observed only for mitochondria isolated from starved and, to a lesser degree, from diabetic rats, and it appeared to result primarily from the enhanced release of approx. 40% and 60%, respectively, of the total CPT activity. This change in the pattern of release was specific to CPT among the marker enzymes studied. For all three types of mitochondria there was no substantial release of CPT concurrently with that of the marker enzyme for the soluble intermembrane space, adenylate kinase. These results illustrate that the bimodal pattern of release of CPT reported previously for mitochondria from starved rats [Bergstrom & Reitz (1980) Arch. Biochem. Biophys. 204, 71-79] is not an immutable consequence of the localization of CPT activity on either side of the mitochondrial inner membrane. Sequential loss of CPT I (i.e. the overt form) from the mitochondrial inner membrane did not affect the concentration of malonyl-CoA required to effect fractional inhibition of the CPT I that remained associated with the mitochondria. The results are discussed in relation to the possibility that altered enzyme-membrane interactions may account for some of the altered regulatory properties of CPT I in liver mitochondria of animals in different physiological states.

scholarly journals Effects of thyroidectomy and starvation on the activity and properties of hepatic carnitine palmitoyltransferase

1982 ◽  
Vol 208 (3) ◽  
pp. 667-672 ◽  
Author(s):  
E D Saggerson ◽  
C A Carpenter ◽  
B S Tselentis
Keyword(s):  
Inhibitory Effect ◽  
Carnitine Palmitoyltransferase ◽  
Hill Coefficient ◽  
Fed State ◽  
Malonyl Coa ◽  
Normal States ◽  
Carnitine Palmitoyltransferase Activity ◽  
The Hill ◽  
Made In

1. Hepatic carnitine palmitoyltransferase activity was measured over a range of concentrations of palmitoyl-CoA and in the presence of several concentrations of the inhibitor malonyl-CoA. These measurements were made in mitochondria obtained from the livers of fed and starved (24 h) normal rats and of fed and starved thyroidectomized rats. 2. In the fed state thyroidectomy substantially decreased overt carnitine palmitoyltransferase activity and also decreased both the Hill coefficient and the s0.5 when palmitoyl-CoA concentration was varied as substrate. Thyroidectomy did not appreciably alter the inhibitory effect of malonyl-CoA on the enzyme. 3. Starvation increased overt carnitine palmitoyltransferase activity in both the fed and the thyroidectomized state. In percentage terms this response to starvation was substantially greater after thyroidectomy. In both the hypothyroid and normal states starvation decreased sensitivity to inhibition by malonyl-CoA.

scholarly journals Response to starvation of hepatic carnitine palmitoyltransferase activity and its regulation by malonyl-CoA. Sex differences and effects of pregnancy

1982 ◽  
Vol 208 (3) ◽  
pp. 673-678 ◽  
Author(s):  
E D Saggerson ◽  
C A Carpenter
Keyword(s):  
Sex Differences ◽  
Virgin Female ◽  
Carnitine Palmitoyltransferase ◽  
Male Rats ◽  
Hill Coefficient ◽  
Acid Oxidation ◽  
Pregnant Rats ◽  
Fed State ◽  
Malonyl Coa ◽  
Carnitine Palmitoyltransferase Activity

1. Hepatic carnitine palmitoyltransferase activity was measured over a range of concentrations of palmitoyl-CoA and in the presence of several concentrations of the inhibitor malonyl-CoA. These measurements were made in mitochondria obtained from the livers of fed and starved (24 h) virgin female and fed and starved pregnant rats. 2. In the fed state overt carnitine palmitoyltransferase activity was significantly lower in virgin females than in age-matched male rats. 3. Starvation increased overt carnitine palmitoyltransferase activity in both virgin and pregnant females. This increase was larger than in the male and was greater in pregnant than in virgin females. 4. In the fed state pregnancy had no effect on the Hill coefficient or the [S]0.5 when palmitoyl-CoA was varied as substrate. Pregnancy did not alter the sensitivity of the enzyme to inhibition by malonyl-CoA. 5. Starvation decreased the sensitivity of the enzyme to malonyl-CoA. The change in sensitivity was similar in male, virgin female and pregnant rats. 6. The possible relevance of these findings to known sex differences and changes with pregnancy in hepatic fatty acid oxidation and esterification are discussed.

scholarly journals Carnitine palmitoyltransferase in human erythrocyte membrane. Properties and malonyl-CoA sensitivity

1991 ◽  
Vol 275 (3) ◽  
pp. 685-688 ◽  
Author(s):  
R R Ramsay ◽  
G Mancinelli ◽  
A Arduini
Keyword(s):  
Membrane Lipids ◽  
Fatty Acyl ◽  
Carnitine Palmitoyltransferase ◽  
Membrane Properties ◽  
Mitochondrial Outer Membrane ◽  
Mitochondrial Inner Membrane ◽  
Malonyl Coa ◽  
Intracellular Enzymes ◽  
Erythrocyte Plasma Membrane ◽  
Triton X

Carnitine palmitoyltransferase located in the erythrocyte plasma membrane is sensitive to inhibition by malonyl-CoA and 2-bromopalmitoyl-CoA plus carnitine. Although this inhibition and other properties suggest similarities to the intracellular enzymes in other tissues, no cross-reaction was observed with antisera to the peroxisomal or to the mitochondrial inner-membrane enzyme. The activity was solubilized by and was stable in Triton X-100, which destroys the enzymes found in microsomes and in the mitochondrial outer membrane. The substrate specificity is broader than for the intracellular enzymes, the activities with stearoyl-CoA (114%) and arachidonoyl-CoA (97%) being equal to that with palmitoyl-CoA, and the activities with linoleoyl-CoA (44%) and erucoyl-CoA (46%) about half that with palmitoyl-CoA. The function of this carnitine palmitoyltransferase is probably to buffer the acyl-CoA present in the erythrocyte for turnover of the fatty acyl groups of the membrane lipids.

scholarly journals Characterization of hepatic carnitine palmitoyltransferase. Use of bromoacyl derivatives and antibodies

1987 ◽  
Vol 241 (3) ◽  
pp. 751-757 ◽  
Author(s):  
P S Brady ◽  
A K Dunker ◽  
L J Brady
Keyword(s):  
Carnitine Palmitoyltransferase ◽  
Dimethyl Sulphoxide ◽  
Inner Membrane ◽  
Phosphatidylcholine Liposomes ◽  
Aqueous Buffer ◽  
Mitochondrial Inner Membrane ◽  
Membrane Enzyme ◽  
Malonyl Coa ◽  
Assay Conditions

Carnitine palmitoyltransferase (CPT) is a mitochondrial-inner-membrane enzyme, with activities located on both the outer and inner sides of the membrane. The inhibition of CPT by bromopalmitate derivatives was studied in intact hepatic mitochondria (representing CPT-A activity, the outer enzyme), in inverted submitochondrial vesicles (representing CPT-B, the inner enzyme), and in purified hepatic CPT. Bromopalmitoyl-CoA had an I50 (concentration giving 50% inhibition of CPT activity) of 0.63 +/- 0.08 microM in intact mitochondria and 2.44 +/- 0.86 microM in inverted vesicles. Preincubation of mitochondria with bromopalmitoyl-CoA decreased V max. for both CPT-A and CPT-B. Sonication decreased sensitivity to bromopalmitoyl-CoA, and solubilization with Triton abolished sensitivity at the concentrations used (0-10 microM). Purified CPT had a bromopalmitoyl-CoA I50 of 353 microM in aqueous buffer, 67 microM in 20% dimethyl sulphoxide, 45 microM in phosphatidylcholine liposomes and 26 microM in cardiolipin liposomes. Increasing [carnitine] at constant bromopalmitoyl-CoA concentrations or increasing [bromopalmitoyl-CoA] in the preincubation resulted in increased inhibition of purified CPT. 2-Tetradecylglycidyl-CoA and malonyl-CoA did not offer measurable protection against bromopalmitoyl-CoA inhibition of the purified CPT, suggesting a different site of interaction of bromopalmitoyl-CoA with CPT. The data suggest that the sensitivity of CPT to bromopalmitoyl-CoA may be modulated by membrane environment and assay conditions.

scholarly journals Interacting effects of l-carnitine and malonyl-CoA on rat liver carnitine palmitoyltransferase

1985 ◽  
Vol 230 (1) ◽  
pp. 161-167 ◽  
Author(s):  
M I Bird ◽  
E D Saggerson
Keyword(s):  
Binding Sites ◽  
Concentration Ratio ◽  
Liver Mitochondria ◽  
Carnitine Palmitoyltransferase ◽  
Heart Mitochondria ◽  
Albumin Concentration ◽  
Latent Form ◽  
Malonyl Coa ◽  
Carnitine Palmitoyltransferase Activity ◽  
Overt Form

Malonyl-CoA significantly increased the Km for L-carnitine of overt carnitine palmitoyltransferase in liver mitochondria from fed rats. This effect was observed when the molar palmitoyl-CoA/albumin concentration ratio was low (0.125-1.0), but not when it was higher (2.0). In the absence of malonyl-CoA, the Km for L-carnitine increased with increasing palmitoyl-CoA/albumin ratios. Malonyl-CoA did not increase the Km for L-carnitine in liver mitochondria from 24h-starved rats or in heart mitochondria from fed animals. The Km for L-carnitine of the latent form of carnitine palmitoyltransferase was 3-4 times that for the overt form of the enzyme. At low ratios of palmitoyl-CoA/albumin (0.5), the concentration of malonyl-CoA causing a 50% inhibition of overt carnitine palmitoyltransferase activity was decreased by 30% when assays with liver mitochondria from fed rats were performed at 100 microM-instead of 400 microM-carnitine. Such a decrease was not observed with liver mitochondria from starved animals. L-Carnitine displaced [14C]malonyl-CoA from liver mitochondrial binding sites. D-Carnitine was without effect. L-Carnitine did not displace [14C]malonyl-CoA from heart mitochondria. It is concluded that, under appropriate conditions, malonyl-CoA may decrease the effectiveness of L-carnitine as a substrate for the enzyme and that L-carnitine may decrease the effectiveness of malonyl-CoA to regulate the enzyme.

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