Control Of Cellular Energy Metabolism

点击翻译按钮获取中文摘要

The purpose of these studies is to establish a better understanding of the energy metabolism of biological tissues using modern system biology approaches. Towards this goal, the laboratory concentrates on the use of screening approaches in proteomics, metabolomics, protein structure, post-translational modifications, minimally invasive metabolic rate information and optical spectroscopy. One of the major hypothesizes in this program is that the activity of the multi-protein Complexes that perform Oxidative Phosphorylation are coordinated in some fashion to balance the rate of ATP production with utilization in the cell. This results in the observed metabolic homeostasis where the potential energy for doing work is maintained near constant in the cell even during major alterations in workload. The following major findings were made over the last year: 1)We have expanded our transmission optical spectroscopy investigation of the functioning of mitochondria in the intact beating heart to include studies on the perfused mouse heart. The small size of the mouse heart required a new optical collection system to detect the light transmitted through the heart with minimal optical artifacts. We developed an approach where the intact heart is placed in the center of an integrating sphere permitting the sampling of all of the light leaving the heart with little or no translational motion artifacts. With Dr. Murphys group we have demonstrated the metabolic state of the mitochondria during ischemia reperfusion conditions and revealed new information on the mitochondrial membrane potential and cytochrome redox state during this clinically relevant protocol. 2) Using exogenous vasodilators in the isolated perfused heart, we have demonstrated the existence of a paradoxical arteriole contraction that puts the isolated heart in partial hypoxia despite the fact that it has flow reserve it is not using. These data implied that the normal regulatory mechanisms of matching blood flow with metabolic demand is disrupted in this major model system of cardiac function. 3) Another vasodilator is vascular nitric oxide, that is also a putative inhibitor of oxidative phosphorylation. We tested the impact of micromolar concentrations of nitric oxide on the isolated perfused rabbit and mouse heart and found no inhibitory action on the cytochrome redox state. We demonstrated that the metabolism of nitric oxide through the oxidation of myoglobin, coupled to a highly active myoglobin reductase, is a powerful nitric oxide metabolizing pathway likely limiting the impact of vascular nitric oxide on mitochondrial function . 3) To broaden our analysis of metabolic regulation in the mitochondria we have expanded our studies to study the ancestors of mitochondria, simple bacteria. We have initiated studies on isolated bacteria believed to be closest to the mitochondrial origins, paracoccus denitrificans(PD). The goal of these studies is to unravel acute energy conversion regulation in this bacterium and then look for similar mechanisms in mammalian mitochondria. With the growing interest in the microbiome, these studies should also provide new insight into the acute regulation of bacterial energy metabolism that has not been extensively studied. We have demonstrated in this period that the previously described metabolic homeostasis described in the mammalian heart, that is constant free energy available in ATP as well as the mitochondria proton motive during increases is workload, exists in PD. This was done with measures of the bacteria membrane potential, transmembrane pH gradient, with a genetically encoded intracellular pH probe, along with the redox state of the cytochromes. We have demonstrated that this property of energy conversion by the cell is conserved over a wide range of biological systems, and hopefully, the bacterial system will provide insights into how the mitochondria were domesticated to perform this task in the human heart and other tissues.