scholarly journals The structural organization of haem synthesis in rat liver mitochondria

1969 ◽  
Vol 113 (3) ◽  
pp. 507-514 ◽  
Author(s):  
M. S. Jones ◽  
O. T. G. Jones
Keyword(s):  
Ultrasonic Treatment ◽  
Mitochondrial Respiratory Chain ◽  
Liver Mitochondria ◽  
Rat Liver Mitochondria ◽  
Outer Membranes ◽  
Soluble Cell ◽  
Cell Components ◽  
Cobalt Protoporphyrin ◽  
Continuous Assay

1. Anaerobic conditions are normally necessary for incorporation of iron into haems and only ferrous iron is used. After addition of succinate to an incubation mixture containing intact or ultrasonically treated mitochondria, Fe3+ is used, but only if no inhibitors prevent the transfer of electrons from the mitochondrial respiratory chain to oxygen. 2. A dual-wavelength spectrophotometric assay for ferrochelatase is described that has been used for the continuous assay of incorporation of metal ions into porphyrins. Constants are given for the determination of rates of formation of protohaem and cobalt protoporphyrin, mesohaem, cobalt mesoporphyrin and zinc mesoporphyrin. For cobalt mesoporphyrin formation the Km for Co2+ is 11×10−6m and that for mesoporphyrin is 5×10−6m. 3. An improved method for the separation of inner and outer membranes of mitochondria is described. Mitochondria swollen in hypo-osmotic media were contracted in hyperosmotic potassium chloride solution containing ATP and the outer membranes detached by mild ultrasonic treatment. Sucrose inhibited the ATP-induced contraction and decreased the yield of outer membranes. 4. Ferrochelatase is associated with cytochrome oxidase, which is used as a marker for inner mitochondrial membranes. 5. By using as substrate porphyrin dissolved in phospholipid micelles, ferrochelatase activity of intact mitochondria was shown to be latent, and to be liberated by ultrasonic treatment. 6. No ferrochelatase was detectable in microsomes or soluble cell components.

scholarly journals Re-evaluation of the interaction of malonyl-CoA with the rat liver mitochondrial carnitine palmitoyltransferase system by using purified outer membranes

1990 ◽  
Vol 267 (1) ◽  
pp. 85-90 ◽  
Author(s):  
M P Kolodziej ◽  
V A Zammit
Keyword(s):  
Rat Liver ◽  
Specific Activity ◽  
Liver Mitochondria ◽  
Subsequent Treatment ◽  
Carnitine Palmitoyltransferase ◽  
Rat Liver Mitochondria ◽  
Outer Membranes ◽  
High Affinity ◽  
Malonyl Coa ◽  
Order Of Magnitude

1. The interaction of malonyl-CoA with the outer carnitine palmitoyltransferase (CPT) system of rat liver mitochondria was re-evaluated by using preparations of highly purified outer membranes, in the light of observations that other subcellular structures that normally contaminate crude mitochondrial preparations also contain malonyl-CoA-sensitive CPT activity. 2. In outer-membrane preparations, which were purified about 200-fold with respect to the inner-membrane-matrix fraction, malonyl-CoA binding was largely accounted for by a single high-affinity component (KD = 0.03 microM), in contrast with the dual site (low- and high-affinity) previously found with intact mitochondria. 3. There was no evidence that the decreased sensitivity of CPT to malonyl-CoA inhibition observed in outer membranes obtained from 48 h-starved rats (compared with those from fed animals) was due to a decreased ratio of malonyl-CoA binding to CPT catalytic moieties. Thus CPT specific activity and maximal high-affinity [14C]malonyl-CoA binding (expressed per mg of protein) were increased 2.2- and 2.0-fold respectively in outer membranes from 48 h-starved rats. 4. Palmitoyl-CoA at a concentration that was saturating for CPT activity (5 microM) decreased the affinity of malonyl-CoA binding by an order of magnitude, but did not alter the maximal binding of [14C]malonyl-CoA. 5. Preincubation of membranes with either tetradecylglycidyl-CoA or 2-bromopalmitoyl-CoA plus carnitine resulted in marked (greater than 80%) inhibition of high-affinity binding, concurrently with greater than 95% inhibition of CPT activity. These treatments also unmasked an effect of subsequent treatment with palmitoyl-CoA to increase low-affinity [14C]malonyl-CoA binding. 6. These data are discussed in relation to the possible mechanism of interaction between the malonyl-CoA-binding site and the active site of the enzyme.

scholarly journals The Influence Of Central Asian Cobra Poison On The Activity Of Rothenon Sensitive And Insensitive Over.H-Oxidase Of Rat Liver Mitochondria And Their Correction With Flavosan (Flateron)

2020 ◽  
Vol 02 (11) ◽  
pp. 96-103
Author(s):  
M. M. Mamajanov ◽  
Keyword(s):  
Respiratory Chain ◽  
Mitochondrial Respiratory Chain ◽  
Liver Mitochondria ◽  
The Body ◽  
Cobra Venom ◽  
Rat Liver Mitochondria ◽  
Inner Mitochondrial Membrane ◽  
Central Asian ◽  
Transport Chain ◽  
Mitochondrial Respiratory

In the presence of cobra venom, the rate of NAD.H oxidation along the internal pathway of the mitochondrial respiratory chain is suppressed, and the rate of NAD.H oxidation along the external pathway increases. These changes occur against the background of cytochrome c deserption from the inner mitochondrial membrane and a significant increase in the process of mitochondrial lipid peroxidation. These facts indicate that when animals are poisoned with cobra venom, profound disturbances are observed in the system of oxidative phosphorylation and the electron transport chain. The introduction of flavosan into the body of animals poisoned with cobra venom leads to an increase in the rate of NAD.H oxidation along the internal pathway of the mitochondrial respiratory chain and suppression of the rate of NAD.H oxidation through the external pathway.

Electron Microscopic Tomography of Whole, Frozen-Hydrated Rat-Liver Mitochondria at 400 kv

1999 ◽  
Vol 5 (S2) ◽  
pp. 416-417
Author(s):  
C.A. Mannella ◽  
C.-E Hsieh ◽  
M. Marko
Keyword(s):  
Rat Liver ◽  
Liver Mitochondria ◽  
Rat Liver Mitochondria ◽  
Peripheral Compartment ◽  
Collection System ◽  
Electron Microscopic ◽  
Automated Data Collection ◽  
Outer Membranes ◽  
Mitochondrial Inner Membrane ◽  
Structural Preservation

Electron microscopic tomography is providing important new insights about the internal structure of the mitochondrion. In particular, the infoldings of the mitochondrial inner membrane (cristae), which are usually rendered as lamelliform baffles, are revealed to have considerable tubular nature. Rather than opening wide to the peripheral compartment (between the inner and outer membranes), the cristae connect to the outside and to each other through narrow (20-30 nm) tubular segments, which can be hundreds of nanometers long. This suggests that diffusion of ions, metabolites and proteins between the intracristal and intermembrane spaces may be restricted.The earlier tomographic reconstructions were done on conventionally prepared, plastic-embedded specimens, which raises the usual concerns about structural preservation. More recently, we have undertaken tomography of isolated rat-liver mitochondria that have been embedded in vitreous ice (by plunge-freezing in iso-osmotic buffer without chemical fixatives or stains). These frozen hydrated specimens are imaged with a JEOL 4000FX equipped with a Gatan cryo-transfer holder and a Tietz automated data collection system, with a Ik × Ik CCD. For 3D reconstructions, images were recorded at a dose of 5 e−Å2 at 2° increments over the range +/− 60°.

scholarly journals Some differences in the properties of carnitine palmitoyltransferase activities of the mitochondrial outer and inner membranes

1987 ◽  
Vol 248 (3) ◽  
pp. 727-733 ◽  
Author(s):  
M S Murthy ◽  
S V Pande
Keyword(s):  
Phospholipase C ◽  
Liver Mitochondria ◽  
Carnitine Palmitoyltransferase ◽  
Mitochondrial Outer Membrane ◽  
Rat Liver Mitochondria ◽  
Outer Membranes ◽  
Mitochondrial Inner Membrane ◽  
Octyl Glucoside ◽  
Malonyl Coa

Recent evidence has shown that the outer, overt, malonyl-CoA-inhibitable carnitine palmitoyltransferase (CPTo) activity resides in the mitochondrial outer membrane [Murthy & Pande (1987) Proc. Natl. Acad. Sci. U.S.A. 84, 378-382]. A comparison of CPTo activity of rat liver mitochondria with the inner, initially latent, carnitine palmitoyltransferase (CPTi) of the mitochondrial inner membrane has revealed that the presence of digitonin and several other detergents inactivates CPTo activity. The CPTi activity, in contrast, was markedly stimulated by various detergents and phospholipid liposomes. These findings explain why in previous studies, which used digitonin or other detergents to expose, separate and purify the CPT activities, the inferences were drawn that (a) the ratio of latent to overt CPT was quite high, (b) both the CPT activities could be ascribed to one active protein recovered, and (c) the observed lack of malonyl-CoA inhibition indicated possible loss/separation of a putative malonyl-CoA-inhibition-conferring protein. Although both CPTo and CPTi were found to catalyse the forward and the backward reactions, CPTo showed greater capacity for the forward reaction and CPTi for the backward reaction. The easily solubilizable CPT, released on sonication of mitoplasts or of intact mitochondria under hypo-osmotic conditions, resembled CPTi in its properties. When octyl glucoside was used under appropriate conditions, 40-50% of the CPTo of outer membranes became solubilized, but it showed limited stability and decreased malonyl-CoA sensitivity. Malonyl-CoA-inhibitability of CPTo was decreased also on exposure of outer membranes to phospholipase C. When outer membranes that had been exposed to octyl glucoside or to phospholipase C were subjected to a reconstitution procedure using asolectin liposomes, the malonyl-CoA-inhibitability of CPTo was restored. A role of phospholipids in the malonyl-CoA sensitivity of CPTo is thus indicated.

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