bims-mimbat 2024-10-27 papers

Issue of 2024‒10‒27
five papers selected by
José Carlos de Lima-Júnior, Washington University

Nat Commun. 2024 Oct 22. 15(1): 9098

Dissected antiporter modules establish minimal proton-conduction elements of the respiratory complex I.

Adel Beghiah, Patricia Saura, Sofia Badolato, Hyunho Kim, Johanna Zipf, Dirk Auman, Ana P Gamiz-Hernandez, Johan Berg, Grant Kemp, Ville R I Kaila.

The respiratory Complex I is a highly intricate redox-driven proton pump that powers oxidative phosphorylation across all domains of life. Yet, despite major efforts in recent decades, its long-range energy transduction principles remain highly debated. We create here minimal proton-conducting membrane modules by engineering and dissecting the key elements of the bacterial Complex I. By combining biophysical, biochemical, and computational experiments, we show that the isolated antiporter-like modules of Complex I comprise all functional elements required for conducting protons across proteoliposome membranes. We find that the rate of proton conduction is controlled by conformational changes of buried ion-pairs that modulate the reaction barriers by electric field effects. The proton conduction is also modulated by bulky residues along the proton channels that are key for establishing a tightly coupled proton pumping machinery in Complex I. Our findings provide direct experimental evidence that the individual antiporter modules are responsible for the proton transport activity of Complex I. On a general level, our findings highlight electrostatic and conformational coupling mechanisms in the modular energy-transduction machinery of Complex I with distinct similarities to other enzymes.

DOI: https://doi.org/10.1038/s41467-024-53194-5

Biochim Biophys Acta Bioenerg. 2024 Oct 21. pii: S0005-2728(24)00488-2. [Epub ahead of print] 149518

Finding the E-channel proton loading sites by calculating the ensemble of protonation microstates.

Md Raihan Uddin, Umesh Khaniya, Chitrak Gupta, Junjun Mao, Gehan A Ranepura, Rongmei Judy Wei, Jose Ortiz-Soto, Abhishek Singharoy, M R Gunner.

The aerobic electron transfer chain builds a proton gradient by proton coupled electron transfer reactions through a series of proteins. Complex I is the first enzyme in the sequence. Here transfer of two electrons from NADH to quinone yields four protons pumped from the membrane N- (negative, higher pH) side to the P- (positive, lower) side. Protons move through three linear antiporter paths, with a few amino acids and waters providing the route; and through the E-channel, a complex of competing paths, with clusters of interconnected protonatable residues. Proton loading sites (PLS) transiently bind protons as they are transported from N- to P-compartments. PLS can be individual residues or extended clusters of residues. The program MCCE uses Monte Carlos sampling to analyze the E-channel proton binding in equilibrium with individual Molecular Dynamics snapshots from trajectories of Thermus thermuphillus Complex I in the apo, quinone and quinol bound states. At pH 7, the five E-channel subunits (Nqo4, Nqo7, Nqo8, Nqo10, and Nqo11) take >25,000 protonation microstates, each with different residues protonated. The microstate explosion is tamed by analyzing interconnected clusters of residues along the proton transfer paths. A proton is bound and released from a cluster of five coupled residues on the protein N-side and to six coupled residues in the protein center. Loaded microstates bind protons to sites closer to the P-side in the forward pumping direction. MCCE microstate analysis identifies strongly coupled proton binding amongst individual residues in the two PLS clusters.

Keywords: Complex I; MCCE; Microstates; Proton pump; Proton transfer

DOI: https://doi.org/10.1016/j.bbabio.2024.149518

EMBO J. 2024 Oct 21.

Glutamine sensing licenses cholesterol synthesis.

Bruna Martins Garcia, Philipp Melchinger, Tania Medeiros, Sebastian Hendrix, Kavan Prabhu, Mauro Corrado, Jenina Kingma, Andrej Gorbatenko, Soni Deshwal, Matteo Veronese, Luca Scorrano, Erika Pearce, Patrick Giavalisco, Noam Zelcer, Lena Pernas.

The mevalonate pathway produces essential lipid metabolites such as cholesterol. Although this pathway is negatively regulated by metabolic intermediates, little is known of the metabolites that positively regulate its activity. We found that the amino acid glutamine is required to activate the mevalonate pathway. Glutamine starvation inhibited cholesterol synthesis and blocked transcription of the mevalonate pathway-even in the presence of glutamine derivatives such as ammonia and α-ketoglutarate. We pinpointed this glutamine-dependent effect to a loss in the ER-to-Golgi trafficking of SCAP that licenses the activation of SREBP2, the major transcriptional regulator of cholesterol synthesis. Both enforced Golgi-to-ER retro-translocation and the expression of a nuclear SREBP2 rescued mevalonate pathway activity during glutamine starvation. In a cell model of impaired mitochondrial respiration in which glutamine uptake is enhanced, SREBP2 activation and cellular cholesterol were increased. Thus, the mevalonate pathway senses and is activated by glutamine at a previously uncharacterized step, and the modulation of glutamine synthesis may be a strategy to regulate cholesterol levels in pathophysiological conditions.

Keywords: Cholesterol; HMGCR; MFN2; Nutrient Sensing; SREBP2

DOI: https://doi.org/10.1038/s44318-024-00269-0

Biochim Biophys Acta Bioenerg. 2024 Oct 18. pii: S0005-2728(24)00489-4. [Epub ahead of print] 149519

How lipid transfer proteins and the mitochondrial membrane shape the kinetics of β-oxidation the liver.

Christoff Odendaal, Dirk-Jan Reijngoud, Barbara M Bakker.

The mitochondrial fatty acid β-oxidation (mFAO) is important for producing ATP under conditions of energetic stress, such as fasting and cold exposure. The regulation of this pathway is dependent on the kinetic properties of the enzymes involved. To better understand pathway behaviour, accurate enzyme kinetics is required. Setting up and interpreting such proper assays requires a good understanding of what influences the enzymes' kinetics. Often, knowing the buffer composition, pH, and temperature is considered to be sufficient. Many mFAO enzymes are membrane-bound, however, and their kinetic properties depend on the composition and curvature of the mitochondrial membranes. These properties are, in turn, affected by metabolite concentrations, but are rarely accounted for in kinetic assays. Especially for carnitine palmitoyltransferase 1 (CPT1), this has been shown to be of great consequence. Moreover, the enzymes of the mFAO metabolise water-insoluble acyl-CoA derivatives, which become toxic at high concentrations. In vivo, these are carried across the cytosol by intracellular lipid transfer proteins (iLTPs), such as the fatty-acid and acyl-CoA-binding proteins (FABP and ACBP, respectively). In vitro, this is often mimicked by using bovine serum albumin (BSA), which differs from the iLPTs in terms of its binding behaviour and subcellular localisation patterns. In this review, we argue that the iLTPs and membrane properties cannot be ignored when measuring or interpreting the kinetics of mFAO enzymes. They should be considered fundamental to the activity of mFAO enzymes just as pH, buffer composition, and temperature are.

DOI: https://doi.org/10.1016/j.bbabio.2024.149519

EMBO J. 2024 Oct 24.

Cyclophilin D plays a critical role in the survival of senescent cells.

Margherita Protasoni, Vanessa López-Polo, Camille Stephan-Otto Attolini, Julian Brandariz, Nicolas Herranz, Joaquin Mateo, Sergio Ruiz, Oscar Fernandez-Capetillo, Marta Kovatcheva, Manuel Serrano.

Senescent cells play a causative role in many diseases, and their elimination is a promising therapeutic strategy. Here, through a genome-wide CRISPR/Cas9 screen, we identify the gene PPIF, encoding the mitochondrial protein cyclophilin D (CypD), as a novel senolytic target. Cyclophilin D promotes the transient opening of the mitochondrial permeability transition pore (mPTP), which serves as a failsafe mechanism for calcium efflux. We show that senescent cells exhibit a high frequency of transient CypD/mPTP opening events, known as 'flickering'. Inhibition of CypD using genetic or pharmacologic tools, including cyclosporin A, leads to the toxic accumulation of mitochondrial Ca2+ and the death of senescent cells. Genetic or pharmacological inhibition of NCLX, another mitochondrial calcium efflux channel, also leads to senolysis, while inhibition of the main Ca2+ influx channel, MCU, prevents senolysis induced by CypD inhibition. We conclude that senescent cells are highly vulnerable to elevated mitochondrial Ca2+ ions, and that transient CypD/mPTP opening is a critical adaptation mechanism for the survival of senescent cells.

Keywords: Cellular Senescence; Cyclophilin D; Mitochondria; Senolytic Therapy; mPTP Flickering

DOI: https://doi.org/10.1038/s44318-024-00259-2