Mitochondria structure, enzymes

Not all the cellular DNA is in the nucleus some is found in the mitochondria. In addition, mitochondria contain RNA as well as several enzymes used for protein synthesis. Interestingly, mitochond-rial RNA and DNA bear a closer resemblance to the nucleic acid of bacterial cells than they do to animal cells. For example, the rather small DNA molecule of the mitochondrion is circular and does not form nucleosomes. Its information is contained in approximately 16,500 nucleotides that func-tion in the synthesis of two ribosomal and 22 transfer RNAs (tRNAs). In addition, mitochondrial DNA codes for the synthesis of 13 proteins, all components of the respiratory chain and the oxidative phosphorylation system. Still, mitochondrial DNA does not contain sufficient information for the synthesis of all mitochondrial proteins most are coded by nuclear genes. Most mitochondrial proteins are synthesized in the cytosol from nuclear-derived messenger RNAs (mRNAs) and then transported into the mito-chondria, where they contribute to both the structural and the functional elements of this organelle. Because mitochondria are inherited cytoplasmically, an individual does not necessarily receive mitochondrial nucleic acid equally from each parent. In fact, mito-chondria are inherited maternally. 

The theory has been advanced that there exists in the cell a particulate structure somewhat smaller than the mitochondrion, the lyso-some, that contains certain autolytic enzymes in a latent situation. The lysosome theory55,56 is very largely based upon measurements made in sucrose homogenates of rodent liver. Although the results for a-D-mannosidase in this tissue (see Table IV) are not incompatible with the theory, the results for other tissues do not always conform to it. In particular, the contrast between mouse and rat spleen argues against a universal single particle to which a-D-mannosidase is confined. Apart from the results quoted in Table IV, not much work has been done on the intracellular location of a-D-mannosidase. 

Vitamin B12 consists of a porphyrin-like ring structure, with an atom of Co chelated at its centre, linked to a nucleotide base, ribose and phosphoric acid (6.34). A number of different groups can be attached to the free ligand site on the cobalt. Cyanocobalamin has -CN at this position and is the commercial and therapeutic form of the vitamin, although the principal dietary forms of B12 are 5 -deoxyadenosylcobalamin (with 5 -deoxyadeno-sine at the R position), methylcobalamin (-CH3) and hydroxocobalamin (-OH). Vitamin B12 acts as a co-factor for methionine synthetase and methylmalonyl CoA mutase. The former enzyme catalyses the transfer of the methyl group of 5-methyl-H4 folate to cobalamin and thence to homocysteine, forming methionine. Methylmalonyl CoA mutase catalyses the conversion of methylmalonyl CoA to succinyl CoA in the mitochondrion. 

In addition to the foregoing more general concerns are questions concerning the localization of an enzyme activity. The location of an enzyme can determine the type of cell lysis, since it could be more advantageous to lyse the cell completely or in such a manner that the organelles are left intact. For example, some lysis methods such as sonication completely disrupt mitochondria, nuclei, and Golgi systems. If an activity is localized in an organelle such as a mitochondrion, it would seem sensible to adopt a method that leaves these structures intact, to facilitate their separation from the rest of the cellular debris. Thus, for the isolation of mitochondrial enzymes, sonication is not the method of choice for cell lysis. 

Protoporphyrinogen oxidase converts protoporphyrinogen IX to the fully desaturated porphyrin in a reaction that uses O2 as the terminal electron acceptor (Fig. 3). The crystal structure of the homodimeric enzyme shows it has one FAD per monomer, which presumably mediates the porphyrin oxidation reaction (19). Like the decarboxylation mediated by coproporphyrinogen oxidase, this reaction also occurs in the mitochondrion. Mutations in the protoporphyrinogen oxidase gene are responsible for variegate porphyria (21). Acute attacks of this disease can be effectively treated by intravenous administration of hematin. 

The synthesis of heme (Fig. 1) is completed in the mitochondrion by insertion of iron into the protoporphyrin IX framework by ferrochelatase. Ferrochelatases from various organisms have been crystallized and their structures determined. The human enzyme contains one 2Fe-2S cluster in each of the two subunits of the functional dimer (22), possibly as a mechanism to link heme synthesis to iron availability. Erythropoietic protoporphyria, which is characterized by cutaneous photosensitivity, is caused by mutations in the ferrochelatase gene (23). 

A quick review of some aspects of mitochondrial structure is in order here because we shall want to describe the exact location of each of the components of the citric acid cycle and the electron transport chain. Recall from Chapter 1 that a mitochondrion has an inner and an outer membrane (Figure 19.2). The region enclosed by the inner membrane is called the mitochondrial matrix, and an intermembrane space exists between the inner and outer membranes. The inner membrane is a tight barrier between the matrix and the cytosol, and very few compounds can cross this barrier without a specific transport protein (Section 8.4). The reactions of the citric acid cycle take place in the matrix, except for the one in which the intermediate electron acceptor is FAD. The enzyme that catalyzes the FAD-linked reaction is an integral part of the inner mitochondrial membrane and is linked direcdy to the electron transport chain (Chapter 20). 

A lipoprotein is a biochemical assembly that contains both proteins and lipids. The lipids or their derivatives may be covalently or noncovalently bound to the proteins. Many enzymes, transporters, structural proteins, antigens, adhesins, and toxins are lipoproteins. Examples include the high density and low density lipoproteins of the blood, the transmembrane proteins of the mitochondrion and the chloroplast, and bacterial lipoproteins [34]. Lipoproteins in the blood, an aqueous medium, carry fats around the body. The protein particles have hydrophilic groups aimed outward so as to attract water molecules this makes them soluble in the salt water based blood pool. Triglyceride-fats and cholesterol are carried internally, shielded from the water by the protein particle [35]. 

Describe the structure of a mitochondrion and identify the location of enzymes important in energy production. 

The first electron carrier in the electron transport chain is an enzyme that contains a tightly bound coenzyme. The coenzyme has a structure similar to FAD. The enzyme formed by the combination of this coenzyme with a protein is called flavin mononucleotide (FMN). Two electrons and one ion from NADH plus another H ion from a mitochondrion pass to FMN, then to an iron-sulfur (Fe—S) protein, and then to coenzyme Q (CoQ). CoQ is also the entry point into the electron transport chain for the two electrons and two H ions from FADH2. As NADH and FADH2 release their hydrogen atoms and electrons, NAD and FAD are regenerated for reuse in the citric acid cycle. 

Fig. 9. Hypothesis on the control of hemoglobin synthesis in chick embryo blastoderm by control of the synthesis of ALA-synthetase. In the nucleus a repressor protein (I) blocks transcription, and a 5jS-H steroid acts as a derepressor, permitting the structural gene (II) to code for the mRNA of ALA-synthetase. In the cytoplasm the information in the mRNA is translated into the enzyme ALA-synthetase (E,) which migrates into the mitochondrion where ALA (III) is made and finally converted by other enzymes (E2-E7) to heme (IV). Heme controls the synthesis of globin either by acting at the initiating site or by permitting proper folding of the globin.

The electrons freed during the oxidation of fatty acids or of the Krebs cycle metabolites are ultimately transferred to the electron transport chain. This transfer can best be achieved if the oxidizing system and the electron transport chain are maintained in close contact in their cellular structures. It is therefore not surprising that, with few exceptions, all the enzymes of the fatty acid oxidation pathway are found in mitochondria. It has not yet been possible to reconstruct the exact pattern of the integration of each of these enzymes within the mitochondrial structure. Available information suggests only that these enzymes are not freely soluble within the mitochondrion. However, they are not as tightly bound to the mitochondrial structure as the enzymes of the electron transport chain. 

Only thiosulfate and thiosulfonates, but not (8-mercaptopyruvate are substrates of rhodanese. Sulfite can replace CN as the S acceptor. The physiological role of rhodanese is unknown, its concentration increases in the fetus until birth, but remains constant in the mother during pregnancy (91). Rhodanese is a latent enzyme in isolated mitochondria and becomes activated upon structural derangement of the mitochondrion caused by aging or hypotonic salt solution (98). 

In view of the increasing number of enzymes whose locale is being pinpointed in differentiated cellular structures, one may well inquire if there is reason to believe that any part of the cell can be considered to be formless in the sense of a homogenous solution. The concept of structure can be extended to the submicroscopic level, and one can conceive of a continuum of structure ranging from the nucleus and mitochondrion at the macro end of the scale to complexes or associations of related enz3nmes at the micro end. 

The association of an enzymatic unit with a structural unit has posed a problem of nomenclature. When one speaks of an enzyme the concept of activity is tacit. An enzyme without activity is no longer an enzyme. However, a mitochondrion as such is recognizable by microscopic and other criteria whether or not it shows enzymatic activity. The term mitochondrion is a structural term and provides no information about the catalytic properties of the mitochrondrion. The term cyclophorase is a functional term and refers to the enzymatic activities exhibited by normal, intact mitochondria. To overcome the difficulty of naming a unit which has both a structural and functional aspect, the term cyclophor-ase-mitochondrial system, abbreviated C.M. system, has been recom-... 

The enzymes connected with fatty acid breakdown also are found in the mitochondria. One structural principle can be derived from the observation that both the formation of active acetate by jS-oxidation and its subsequent consumption in the citrate cycle take place in the same subcellular particle, the mitochondrion. 

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