Proposed proton pumping mechanism in Mitochondria; How to Mitochondria pump protons to regions of high proton concentration?

Author: Dr. Sang-Tae, Lee

Presented on 29^th June 2023

Mitochondria, the powerhouses of the cell, play a crucial role in generating ATP through oxidative phosphorylation. One of the key processes in this energy production is the pumping of proton (H⁺) to region of higher concentration within the mitochondria.

Cytochrome c oxidase (CcO) is an enzyme located on the inner membrane of mitochondria. It facilitates the reduction of oxygen (O₂) to water (H₂O) through the following reaction; O₂ + 8H⁺ + 4e^- à 2 H₂O + 4 H⁺ + heat

The energy generated during this process is utilized to pump four protons across the intermembrane space. The resulting high concentration of protons creates a gradient that drives the synthesis of ATP in the mitochondrial matrix, Figure 1.

Figure 1. Mitochondria’s working

The role of CcO in Complex IV is to provide electrons to oxygen for reduction to water. However, there is still something that is not explained. How can hydrogen ions be pumped from the mitochondrial matrix, where the concentration of hydrogen ions is low, to the inter membrane, where the concentration of hydrogen ions is high? I have been studying CcO modelling for a long time, and here I would like to explain the mechanism with a new idea.

The proton pumping mechanism is proposed as shown in Figure 2. Let us assume that the left inter membrane beaker is a beaker at 25 degrees with a high concentration of hydrogen ions, and the right mitochondrial matrix beaker with a low concentration of hydrogen ions is connected by an intermediate valve. Normally, when the valve is opened, hydrogen ions will diffuse down from the high-concentration hydrogen ion side. However, if the mitochondrial matrix beaker with a low concentration of hydrogen ions is heated and the temperature is higher than 25 degrees, the situation changes. According to the Fick's the first law of diffusion (J = -DΔC + kΔT), the movement of fluids is affected by the concentration difference and the temperature difference, but if the temperature difference is large, the material at a higher temperature will move to a lower temperature. If it is left alone, it will reach parallel, but if the valve is opened and closed, hydrogen ions will be pumped from the high-temperature side to the low-temperature side even if the concentration of hydrogen ions is low.

Figure 2. The diagram shows the proposed mechanism for proton pumping in CcO.

To explain this natural phenomenon, there must be devices inside CcO that can explain the above. One is a lamp that emits heat continuously, and the other is a chemical structure that acts as a valve. First, the lamp is the reaction of oxygen being reduced between copper and iron, as shown in Figure 3. This reaction is an exothermic reaction that will continuously raise the temperature of the surrounding water, and this will cause a temperature difference between the mitochondrial matrix and the inter membrane. Second, according to the journal paper1 published by Steve and Sang, Histine crosslinked Tyrosine (His-Tyr) is a compound that is bound to copper in a different protein, unlike two Histines groups connected to a single protein, Figure 3.

Figure 3. The diagram shows the chemical structure of cytochrome c oxidase and the proposed mechanism for proton pumping with corresponding chemical structure.

Here, His-Tyr is a compound that receives electrons from the outside and provides them to the Copper center. At this time, valence tautomerism occurs between Cu(II)-His-TyrO^- and Cu(I) - - His-TyrO*, and the Cu-Histidine bond is temporarily broken when the Cu is in the Cu(I) state (the latter). Therefore, His-Tyr acts as a valve in the above model. The bound Cu(II)-His-TyrO^- is a closed valve, and the unbound Cu(I)- - His-TyrO* is an open valve.

When oxygen is reduced at the CcO center, heat is generated, and protons are pumped from the mitochondrial matrix, where the concentration of proton ions is low, to the inter membrane, where the concentration of proton ions is high. His-Tyr acts as a valve to prevent thermal equilibrium and proton concentration equilibrium.

Reference;

1. Am. Chem. Soc. 2007, 129, 5800-5801 Valence Tautomerism and Coordinative Lability in Copper (II)-Imidazoly-Semiquiononate Anion Radical Models for the Cu_B Centre in Cytochrome c Oxidases.

Author: Dr. Sang-Tae, Lee

2005 PhD Bio-inorganic Chemistry UNSW Australia

Contact: sangtael@gmail.com