Respiration of mitochondria in living organisms is controlled by the ATP/ADP ratio and phosphorylation pattern of cytochrome c oxidase

The "second mechanism of respiratory control" (allosteric ATP-inhibition of cytochrome c oxidase (CcO)) is demonstrated for the first time in intact isolated rat liver and heart mitochondria. The problems of measuring the kinetics of allosteric ATP-inhibition in isolated mitochondria were investigated. And it was found that only at very high ATP/ADP ratios, this inhibition is obtained and requires an ATP-regenerating system consisting of phosphoenolpyruvate (PEP) and pyruvate kinase (PK). The allosteric ATP-inhibition can be switched off probably by dephosphorylation of a serine at CcO subunit-I. The phosphorylation of CcO at serine, threonine and tyrosine was studied in isolated mitochondria by extracting complex IV of the respiratory chain (CcO) by BN-PAGE (blue-nativepolyacrylamide- gel-electrophoresis), SDS-PAGE and Western blotting with the corresponding antibodies against the phosphorylated amino acids. The extent of allosteric ATP-inhibition of CcO varied in different preparations of mitochondria, which was suggested to be based on the psychological situation of the animal at the time of killing. Incubation of bovine heart tissue slices with insulin decreased the allosteric ATP-inhibition and phosphorylation of CcO subunit-I at serine. By using a TPP+ (tetraphenylphosphonium)- electrode, and in collaboration with Katrin Staniek, Veterinary University, Vienna, a reversible decrease of the mitochondrial membrane potential (DYm) by PEP and PK through the DYm-independent allosteric ATP-inhibition of CcO at high ATP/ADP ratios was demonstrated in isolated rat liver mitochondria for the first time. It is proposed that respiration in living eukaryotic organisms is normally regulated by the DYm-independent ‘allosteric ATPinhibition of CcO’, and only when the allosteric ATP-inhibition is switched off under stress, respiration is regulated by ‘respiratory control’, based on DYm according to the Mitchell Theory

The ABCT31 Transporter Regulates the Export System of Phenylacetic Acid as a Side-Chain Precursor of Penicillin G in Monascus ruber M7

The biosynthesis of penicillin G (PG) is compartmentalized, and the transportation of the end and intermediate products, and substrates (precursors) such as L-cysteine (L-Cys), L-valine (L-Val) and phenylacetic acid (PAA) requires traversing membrane barriers. However, the transportation system of PAA as a side chain of PG are unclear yet. To discover ABC transporters (ABCTs) involved in the transportation of PAA, the expression levels of 38 ABCT genes in the genome of Monascus ruber M7, culturing with and without PAA, were examined, and found that one abct gene, namely abct31, was considerably up-regulated with PAA, indicating that abct31 may be relative with PAA transportation. Furthermore the disruption of abct31 was carried out, and the effects of two PG substrate's amino acids (L-Cys and L-Val), PAA and some other weak acids on the morphologies and production of secondary metabolites (SMs) of Δabct31 and M. ruber M7, were performed through feeding experiments. The results revealed that L-Cys, L-Val and PAA substantially impacted the morphologies and SMs production of Δabct31 and M. ruber M7. The UPLC-MS/MS analysis findings demonstrated that Δabct31 did not interrupt the synthesis of PG in M. ruber M7. According to the results, it suggests that abct31 is involved in the resistance and detoxification of the weak acids, including the PAA in M. ruber M7

Comparative Assessment of the Bioremedial Potentials of Potato Resistant Starch-Based Microencapsulated and Non-encapsulated Lactobacillus plantarum to Alleviate the Effects of Chronic Lead Toxicity

Lead (Pb) is a well-recognized and potent heavy metal with non-biodegradable nature and can induce the oxidative stress, degenerative damages in tissues, and neural disorders. Certain lactic acid bacterial strains retain the potential to mitigate the lethal effects of Pb. The present work was carried out to assess the Pb bio-sorption and tolerance capabilities of Lactobacillus plantarum spp. Furthermore, potato resistant starch (PRS)-based microencapsulated and non-encapsulated L. plantarum KLDS 1.0344 was utilized for bioremediation against induced chronic Pb toxicity in mice. The experimental mice were divided into two main groups (Pb exposed and non-Pb exposed) and, each group was subsequently divided into three sub groups. The Pb exposed group was exposed to 100 mg/L Pb(NO3)2 via drinking water, and non-Pb exposed group was supplied with plain drinking water during 7 weeks prolonged in vivo study. The accumulation of Pb in blood, feces, renal, and hepatic tissues and its pathological damages were analyzed. The effect of Pb toxicity on the antioxidant enzyme capabilities in blood, serum, as well as, on levels of essential elements in tissues was also calculated. Moreover, KLDS 1.0344 displayed remarkable Pb binding capacity 72.34% and Pb tolerance (680 mg/L). Oral administration of both non- and PRS- encapsulated KLDS 1.0344 significantly provided protection against induced chronic Pb toxicity by increasing fecal Pb levels (445.65 ± 22.28 μg/g) and decreasing Pb in the blood up to 137.63 ± 2.43 μg/L, respectively. KLDS 1.0344 microencapsulated with PRS also relieved the renal and hepatic pathological damages and improved the antioxidant index by inhibiting changes in concentrations of glutathione peroxidase, glutathione, superoxide dismutase, malondialdehyde, and activated oxygen species, which were affected by the Pb exposure. Overall, our results suggested that L. plantarum KLDS 1.0344 either in free or encapsulated forms hold the potentiality to deliver a dietetic stratagem against Pb lethality

Cytochrome c Oxidase Inhibition by ATP Decreases Mitochondrial ROS Production

This study addresses the eventual consequence of cytochrome c oxidase (CytOx) inhibition by ATP at high ATP/ADP ratio in isolated rat heart mitochondria. Earlier, it has been demonstrated that the mechanism of allosteric ATP inhibition of CytOx is one of the key regulations of mitochondrial functions. It is relevant that aiming to maintain a high ATP/ADP ratio for the measurement of CytOx activity effectuating the enzymatic inhibition as well as mitochondrial respiration, optimal concentration of mitochondria is critically important. Likewise, only at this concentration, were the differences in ΔΨ(m) and ROS concentrations measured under various conditions significant. Moreover, when CytOx activity was inhibited in the presence of ATP, mitochondrial respiration and ΔΨ(m) both remained static, while the ROS production was markedly decreased. Consubstantial results were found when the electron transport chain was inhibited by antimycin A, letting only CytOx remain functional to support the energy production. This seems to corroborate that the decrease in mitochondrial ROS production is solely the effect of ATP binding to CytOx which results in static respiration as well as membrane potential

Confirmation of root-knot nematode resistant gene Rmi1 using SSR markers

Background: The Root Knot Nematode (RKN) is a serious economic threat to various cultivated crops worldwide. It is a devastating pest of soybean and responsible to cause severe yield loss in Pakistan. The cultivation of resistant soybean varieties against this pest is the sustainable strategy to manage the heavy loss and increase yield. There is an utmost need to identify RKN resistant varieties of soybean against cultivated in Pakistan. The presented study is an attempt to identify and confirm the presence of resistant gene Rmi1 in soybean. Method: Molecular studies have been done using Simple Sequence Repeat (SSR) marker system to identify resistant soybean varieties against Root Knot Nematode (RKN) using fifteen (15) indigenous cultivars and four (4) US cultivars. DNA was isolated, purified, quantified and then used to employ various SSR markers. The amplified product is observed using gel documentation system after electrophoresis.  Results: Diagnostic SSR markers Satt-358 and Satt-492 have shown the presence of Rmi1 gene in all resistance carrying genotypes. Satt-358 amplified the fragment of 200 bp and Satt-492 generated 232 bp bands in all resistant genotypes. This study confirmed the Rmi gene locus (G248A-1) in all internationally confirmed resistant including six (6) native varieties.Conclusion: These investigations have identified six (6) resistant cultivars revealing the effective and informative sources that can be utilized in breeding programs for the selection of RKN resistance soybean genotypes in Pakistan.

Mitochondrial physiology

As the knowledge base and importance of mitochondrial physiology to evolution, health and disease expands, the necessity for harmonizing the terminology concerning mitochondrial respiratory states and rates has become increasingly apparent. The chemiosmotic theory establishes the mechanism of energy transformation and coupling in oxidative phosphorylation. The unifying concept of the protonmotive force provides the framework for developing a consistent theoretical foundation of mitochondrial physiology and bioenergetics. We follow the latest SI guidelines and those of the International Union of Pure and Applied Chemistry (IUPAC) on terminology in physical chemistry, extended by considerations of open systems and thermodynamics of irreversible processes. The concept-driven constructive terminology incorporates the meaning of each quantity and aligns concepts and symbols with the nomenclature of classical bioenergetics. We endeavour to provide a balanced view of mitochondrial respiratory control and a critical discussion on reporting data of mitochondrial respiration in terms of metabolic flows and fluxes. Uniform standards for evaluation of respiratory states and rates will ultimately contribute to reproducibility between laboratories and thus support the development of data repositories of mitochondrial respiratory function in species, tissues, and cells. Clarity of concept and consistency of nomenclature facilitate effective transdisciplinary communication, education, and ultimately further discovery

Mitochondrial physiology

As the knowledge base and importance of mitochondrial physiology to evolution, health and disease expands, the necessity for harmonizing the terminology concerning mitochondrial respiratory states and rates has become increasingly apparent. The chemiosmotic theory establishes the mechanism of energy transformation and coupling in oxidative phosphorylation. The unifying concept of the protonmotive force provides the framework for developing a consistent theoretical foundation of mitochondrial physiology and bioenergetics. We follow the latest SI guidelines and those of the International Union of Pure and Applied Chemistry (IUPAC) on terminology in physical chemistry, extended by considerations of open systems and thermodynamics of irreversible processes. The concept-driven constructive terminology incorporates the meaning of each quantity and aligns concepts and symbols with the nomenclature of classical bioenergetics. We endeavour to provide a balanced view of mitochondrial respiratory control and a critical discussion on reporting data of mitochondrial respiration in terms of metabolic flows and fluxes. Uniform standards for evaluation of respiratory states and rates will ultimately contribute to reproducibility between laboratories and thus support the development of data repositories of mitochondrial respiratory function in species, tissues, and cells. Clarity of concept and consistency of nomenclature facilitate effective transdisciplinary communication, education, and ultimately further discovery

Multiple Mechanisms Regulate Eukaryotic Cytochrome C Oxidase

Cytochrome c oxidase (COX), the rate-limiting enzyme of mitochondrial respiration, is regulated by various mechanisms. Its regulation by ATP (adenosine triphosphate) appears of particular importance, since it evolved early during evolution and is still found in cyanobacteria, but not in other bacteria. Therefore the “allosteric ATP inhibition of COX” is described here in more detail. Most regulatory properties of COX are related to “supernumerary” subunits, which are largely absent in bacterial COX. The “allosteric ATP inhibition of COX” was also recently described in intact isolated rat heart mitochondria

Multiple Mechanisms Regulate Eukaryotic Cytochrome C Oxidase

Cytochrome c oxidase (COX), the rate-limiting enzyme of mitochondrial respiration, is regulated by various mechanisms. Its regulation by ATP (adenosine triphosphate) appears of particular importance, since it evolved early during evolution and is still found in cyanobacteria, but not in other bacteria. Therefore the “allosteric ATP inhibition of COX” is described here in more detail. Most regulatory properties of COX are related to “supernumerary” subunits, which are largely absent in bacterial COX. The “allosteric ATP inhibition of COX” was also recently described in intact isolated rat heart mitochondria

Stress-mediated generation of deleterious ROS in healthy individuals - role of cytochrome c oxidase

Psychosocial stress is known to cause an increased incidence of coronary heart disease. In addition, multiple other diseases like cancer and diabetes mellitus have been related to stress and are mainly based on excessive formation of reactive oxygen species (ROS) in mitochondria. The molecular interactions between stress and ROS, however, are still unknown. Here we describe the missing molecular link between stress and an increased cellular ROS, based on the regulation of cytochrome c oxidase (COX). In normal healthy cells, the 'allosteric ATP inhibition of COX' decreases the oxygen uptake of mitochondria at high ATP/ADP ratios and keeps the mitochondrial membrane potential (Δ