Substrate and oxidative phosphorylation

Substrate-level phosphorylation is a type of chemical reaction that results in the formation and creation of adenosine triphosphate (ATP) by the direct transfer and donation of a phosphoryl (PO3) group to adenosine diphosphate (ADP) from a reactive intermediate. While technically the transfer is PO3, or a phosphoryl group, convention in biological sciences is to refer to this as the transfer of a phosphate group. In cells, it occurs primarily and firstly in the cytoplasm (in glycolysis) under both aerobic and anaerobic conditions.

Unlike oxidative phosphorylation, here the oxidation and phosphorylation are not coupled or joined, although both types of phosphorylation result in ATP. It should be noted that there is an oxidation reaction coupled to phosphorylation, however this occurs in the generation of 1,3- bisphosphoglycerate from 3- phosphoglyceraldehyde via a dehydrogenase. ATP is generated in a separate step (key difference from oxidative phosphorylation) by transfer of the high energy phosphate on 1,3-bisphosphoglycerate to ADP via a kinase.

ATP is synthesized when protons flow back to the mitochondrial matrix through an enzyme complex ATP synthase. The oxidation of fuels and the phosphorylation of ADP are coupled by a proton gradient across the inner mitochondrial membrane. Oxidative phosphorylation is the process in which ATP is formed as a result of the transfer of electrons from NADH or FADH 2 to O 2 by a series of electron carriers.

OXIDATIVE PHOSPHORYLATION IN EUKARYOTES TAKES PLACE IN MITOCHONDRIA Two membranes: outer membrane inner membrane (folded into cristae) Two compartments: (1) the intermembrane space (2) the matrix Inner mitochondrial membrane: Electron transport chain ATP synthase Mitochondrial matrix: Pyruvate dehydrogenase complex Citric acid cycle Fatty acid oxidation Location of mitochondrial complexes The outer membrane is permeable to small molecules and ions because it contains pore-forming protein (porin). The inner membrane is impermeable to ions and polar molecules. Contains transporters (translocases).

THE ELECTRON TRANSPORT CHAIN Series of enzyme complexes (electron carriers) embedded in the inner mitochondrial membrane, which oxidize NADH 2 and FADH 2 and transport electrons to oxygen is called respiratory electron-transport chain (ETC). The sequence of electron carriers in ETC cyt b NADH FMN Fe-S Co-Q Fe-S cyt c 1 cyt c cyt a cyt a 3 O 2 succinate FAD Fe-S

High-Energy Electrons: Redox Potentials and Free-Energy Changes In oxidative phosphorylation, the electron transfer potential of NADH or FADH 2 is converted into the phosphoryl transfer potential of ATP. Phosphoryl transfer potential is  G°' (energy released during the hydrolysis of activated phos- phate compound).  G°' for ATP = -7.3 kcal mol -1 Electron transfer potential is expressed as E' o, the (also called redox potential, reduction potential, or oxidation-reduction potential).

E' o (reduction potential) is a measure of how easily a compound can be reduced (how easily it can accept electron). All compounds are compared to reduction potential of hydrogen wich is 0.0 V. The larger the value of E' o of a carrier in ETC the better it functions as an electron acceptor (oxidizing factor). Electrons flow through the ETC components spontaneously in the direction of increasing reduction potentials. E' o of NADH = volts (strong reducing agent) E' o of O 2 = volts (strong oxidizing agent) cyt b NADH FMN Fe-S Co-Q Fe-S cyt c 1 cyt c cyt a cyt a 3 O 2 succinate FAD Fe-S

Important characteristic of ETC is the amount of energy released upon electron transfer from one carrier to another. This energy can be calculated using the formula:  G o ’=-nF  E’ o n – number of electrons transferred from one carrier to another; F – the Faraday constant (23.06 kcal/volt mol);  E’ o – the difference in reduction potential between two carriers. When two electrons pass from NADH to O 2 :  G o ’=-2*96,5*(+0,82-(-0,32)) = kcal/mol