Proline Oxidation Supports Mitochondrial ATP Production When Complex I Is Inhibited

Abstract

The oxidation of proline to pyrroline-5-carboxylate (P5C) leads to the transfer of electrons to ubiquinone in mitochondria that express proline dehydrogenase (ProDH). This electron transfer supports Complexes CIII and CIV, thus generating the protonmotive force. Further catabolism of P5C forms glutamate, which fuels the citric acid cycle that yields the reducing equivalents that sustain oxidative phosphorylation. However, P5C and glutamate catabolism depend on CI activity due to NAD+ requirements. Next, Gen-O2k (Oroboros Instruments) was used to measure proline oxidation in isolated mitochondria of various mouse tissues. Simultaneous measurements of oxygen consumption, membrane potential, NADH, and the ubiquinone redox state were correlated to ProDH activity and F1FO-ATPase directionality. Proline catabolism generated a sufficiently high membrane potential that was able to maintain the F1FO-ATPase operation in the forward mode. This was observed in CI-inhibited mouse liver and kidney mitochondria that exhibited high levels of proline oxidation and ProDH activity. This action was not observed under anoxia or when either CIII or CIV were inhibited. The duroquinone fueling of CIII and CIV partially reproduced the effects of proline. Excess glutamate, however, could not reproduce the proline effect, suggesting that processes upstream of the glutamate conversion from proline were involved. The ProDH inhibitors tetrahydro-2-furoic acid and, to a lesser extent, S-5-oxo-2-tetrahydrofurancarboxylic acid abolished all proline effects. The data show that ProDH-directed proline catabolism could generate sufficient CIII and CIV proton pumping, thus supporting ATP production by the F1FO-ATPase even under CI inhibition.

Improved cardioprotective effect of 3‐nitro‐N‐methyl salicylamide solution after a prolonged preservation time of rat heart

“…The purified mitochondrial solution containing 300 μg protein was added to the chamber for detection. Oxygen concentration (μM) and oxygen flux (pmol‐s‐mg‐1; negative time derivative of oxygen concentration divided by mitochondrial mass per volume and corrected for instrumental background oxygen flux resulting from oxygen consumption of the oxygen sensor and back‐diffusion into the chamber) were recorded using DatLab 5.0 software (Oroboros Instruments, Austria) 18 . The RCR was expressed as state3/state4 respiratory rate.…”

Proline Maintains the Proliferation of Hepatocellular Carcinoma Cells by Decreasing Intracellular Oxidative Stress and Reducing Autophagy During Acute Nutrient Stress

“…The PYCR1 and ALDH18A1 genes, which regulate proline synthesis, are frequently highly expressed in tumour cells and contribute to proliferation and invasion [14]. Retrospective analysis has shown that proline metabolism is associated with the progression of breast, kidney cancer, colorectal, and liver cancer, as well as melanoma [15][16][17]. Elia et al [18] reported that proline metabolism increased cancer stemness and supported metastasis in breast cancer.…”

A method to assess the mitochondrial respiratory capacity of complexes I and II from frozen tissue using the Oroboros O2k-FluoRespirometer

“…However, they are most widely studied for the capacity to produce the biological energy currency of ATP, through the coupling of oxidative phosphorylation with the electron transfer system. The electron transfer system is composed of a series of enzyme complexes in the inner mitochondrial membrane that oxidise a range of metabolites, including NADH derived from the TCA cycle 5 , fatty acids via beta-oxidation 6 , succinate 7 , proline 8 , and glycerol-3-phosphate in mammals 9 . Electrons derived from these species are funnelled into the Q-junction (co-enzyme Q), before the sequential movement through complex III, cytochrome c and finally respiratory complex IV, where they reduce molecular oxygen in a system that has been well-reviewed 10 .…”