Biological Oxidation and Oxidative Phosphorylation of Oxidative Phosphorylates

In the process of transferring high-potential electrons from NADH or FADH2 to oxygen along the respiratory chain, the released energy is transferred to ADP to form ATP, that is, the formation of ATP is coupled with electron transfer, which is called oxidative phosphorylation, and its characteristic is that oxygen molecules need to participate.

Oxidative phosphorylation is different from substrate-level phosphorylation: substrate-level phosphorylation refers to the redistribution of energy in metabolic substrate due to dehydrogenation or dehydration, and the energy carried by the generated high-energy bonds is transferred to ADP to generate ATP, that is, the formation of ATP is directly coupled with the transfer of phosphate groups on a high-energy phosphate compound in metabolism (such as phosphoenolpyruvate, 1, 3- diphosphoglyceric acid, etc.). ), which is characterized by no molecular oxygen. Decoupling of (1) oxidative phosphorylation

In intact mitochondria, electron transfer and phosphorylation are closely coupled. When the two processes of electron transfer and ATP formation are separated by some reagents, only electron transfer cannot form ATP, which is called decoupling.

(2) Decoupling agent for oxidative phosphorylation

The reagent that can cause decoupling is called decoupling agent. The essence of decoupling is that the decoupling agent eliminates the transmembrane proton concentration or potential gradient produced by electron transfer, only electron transfer, no ATP.

(3) Type of decoupling agent

A typical decoupling agent is chemical 2,4-dinitrophenol (DNP), which is weakly acidic and can bind or release H in different pH environments. Moreover, DNP is fat-soluble and can penetrate phospholipid bilayer, so that H can be transferred from the outer side to the inner side of mitochondrial intima, thus eliminating the H gradient. In addition, ionophores, such as valinomycin, an antibiotic produced by streptomycin, are fat-soluble and can coordinate with K ions, so that K outside the mitochondrial membrane can be transported to the membrane and the transmembrane potential gradient can be eliminated. In addition, there is a natural uncoupling protein in the mitochondrial inner membrane of some biological cells. The proton channel formed by protein can make the protons outside the membrane return to the membrane through the channel, so as to eliminate the proton concentration gradient across the membrane, but it cannot generate ATP and heat to raise the body temperature.

Decoupling agents are different from electron transfer inhibitors. Decoupling agent only eliminates the proton or potential gradient on both sides of the intima, does not inhibit the electron transfer in the respiratory chain, and even accelerates the electron transfer, promoting the consumption of respiratory substrates and molecular oxygen, but does not form ATP, only generates heat. Although the mechanism of oxidative phosphorylation and electron transfer coupling has been studied for many years, it is still unclear. There are three hypotheses trying to explain its mechanism. These three hypotheses are: chemical coupling hypothesis, conformational coupling hypothesis and chemical permeation hypothesis.

(1) chemical coupling hypothesis

It is considered that the free energy released in electron transfer temporarily exists in the form of high-energy valence intermediate, and then it is cracked to transfer its energy to ADP to form ATP. But we can't find examples of high-energy intermediates in respiratory chain.

(2) Conformational coupling hypothesis

It is considered that the free energy released by electrons along the respiratory chain causes the conformational changes of mitochondrial inner membrane proteins, forming high-energy forms and temporarily existing. This high-energy form transfers energy to F0F 1-ATPase molecule, changing its conformation. When f0f1-ATPase is recovered, energy is transferred to ADP to form ATP.

(3) chemical infiltration hypothesis

This hypothesis was put forward by British biochemist Peter Mitchell. He believes that the result of electron transfer is to "pump" H from the inner side of the mitochondrial intima to the outer side of the intima, thus generating a concentration gradient of H on the inner and outer sides of the intima. That is, there is potential energy between the outer and inner intima, which is the driving force for H to return to the inner intima. H flows back to the inner side of the intima through special channels on F0F 1-ATPase molecules. When H returns to the inner side of intima, the reaction of releasing free energy is coupled with ATP synthesis reaction. This assumption has been supported by many people at present.

Experiments show that oxidative phosphorylation requires a complete mitochondrial inner membrane. When the mitochondria are treated with ultrasound, the crista intima of the mitochondria can be broken into fragments: the crista membrane of some fragments is closed again to form vesicles, which are called lower mitochondrial vesicles (inner membrane is everted). These sub-mitochondrial vesicles still have the function of oxidative phosphorylation. F 1 spheroid can be seen outside the vesicle. When these vesicles are treated with urea or trypsin, the ball F 1 on the intima falls off and F0 remains on the membrane. The treated vesicles still have the function of electron transport chain, but lose the function of ATP synthesis. When the spheroid of F 1 was added back to the vesicle with only F0, oxidative phosphorylation resumed. This experiment shows that the enzyme (F0) on the inner crista of mitochondria plays the role of electron transfer, and the F 1 on it is an important component to form ATP. F0 and F 1 are enzyme complexes. 1, quantitative measurement

There are three kinds of adenylate in cells, namely AMP, ADP and ATP, which are called adenylate pool. The relative amounts of ATP, ADP and AMP in a cell at a certain time control the activity of the cell. Atkinson put forward the concept of energy charge. It is considered that the energy charge is a quantitative measure of the high-energy phosphate state of cells, and the energy charge can explain the energy state of ATP-ADP-AMP system in organisms.

2. Determinants of energy charge.

It can be seen that the energy charge depends on the amount of ATP and ADP. The energy charge ranges from 0 to 1.0. When all cells are ATP, the energy charge is 1.0. At this time, the number of available high-energy phosphate bonds is the largest. When they are all ADP, the energy charge is 0.5, and there are half high-energy phosphate bonds in the system. When they are all AMP, the energy charge is 0, and there is no high-energy phosphoric acid compound at this time. Experiments show that high-energy charge can inhibit the production of ATP, but promote the utilization of ATP. In other words, high-energy charges can promote anabolism and inhibit catabolism, whereas low-energy charges can promote catabolism and inhibit catabolism.

The energy charge regulation is realized by the allosteric regulation of ATP, ADP and AMP molecules on some enzyme molecules. NADH produced by glycolysis in the cytoplasm of eukaryotes cannot be oxidized directly through the inner membrane of mitochondria, but protons on NADH+H can enter the electron transfer chain indirectly through shuttle. The shuttle process of glycerol 3- phosphate was first discovered. The process is that NADH- 1H in cytoplasm reacts with dihydroxyacetone phosphate under the action of glycerol 3- phosphate dehydrogenase to produce glycerol 3- phosphate. Glycerol-3- phosphate can enter mitochondria, and under the action of glycerol-3- phosphate dehydrogenase (FAD as auxiliary group) on the inner membrane of mitochondria, dihydroxyacetone phosphate and FADH2 are generated. Dihydroxyacetone phosphate permeates mitochondria and continues to act as a hydrogen acceptor. FADH2 transfers hydrogen to CoQ and enters respiratory chain for oxidation, which can only produce ATP at two points.

There is another shuttle mode in mitochondria of animal liver, kidney and heart, namely oxaloacetate-malic acid shuttle. In this way, the dehydrogenase coenzyme in cytosol and mitochondria is NAD+, so NADH in cytosol reaches mitochondria and generates NADH+H. From the point of energy production, the oxaloacetate-malic acid shuttle mechanism is better than α -glycerophosphate shuttle mechanism. However, the shuttle mechanism of α -glycerophosphate is much faster than that of oxaloacetate-malic acid.