BACKGROUND: Mitochondrial energy can be stored as ATP or released as heat by uncoupling protein 1 (UCP1) during non-shivering thermogenesis in brown adipose tissue. UCP1, located in the inner mitochondrial membrane, reduces the proton gradient in the presence of long-chain fatty acids (FA). FA act as weak, protein-independent uncouplers, with the transport of the FA anion across the membrane being the rate-limiting step. According to the fatty acid cycling hypothesis, UCP1 catalyzes this step through an as-yet-undefined mechanism. METHODS: We used computational and experimental techniques, including all-atom molecular dynamics (MD) simulations, membrane conductance measurements, and site-directed mutagenesis. RESULTS: We identified two novel pathways for fatty acid anion translocation (sliding) at the UCP1 protein-lipid interface, ending at key arginine residues R84 and R183 in a nucleotide-binding region. This region forms a stable complex with fatty acid anion, which is crucial for anion transport. Mutations of these two arginines reduced membrane conductance, consistent with the MD simulation prediction that the arachidonic acid anion slides between helices H2-H3 and H4-H5, terminating at R84 and R183. Protonation of the arachidonic acid anion predicts its release from the protein-lipid interface, allowing it to move to either cytosolic or matrix leaflets of the membrane. CONCLUSION: We provide a novel, detailed mechanism by which UCP1 facilitates fatty acid anion transport, as part of the fatty acid cycling process originally proposed by Skulachev. The residues involved in this transport are conserved in other SLC25 proteins, suggesting the mechanism may extend beyond UCP1 to other members of the superfamily.
- Klíčová slova
- anion transporter, cardiolipin, fatty acid cycling, mitochondrial SLC25 family, molecular dynamics simulations, proton transport mechanism,
- MeSH
- iontové kanály * metabolismus chemie genetika MeSH
- iontový transport fyziologie MeSH
- lidé MeSH
- mastné kyseliny * metabolismus MeSH
- mitochondriální proteiny * metabolismus chemie genetika MeSH
- mutageneze cílená MeSH
- simulace molekulární dynamiky * MeSH
- uncoupling protein 1 MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- iontové kanály * MeSH
- mastné kyseliny * MeSH
- mitochondriální proteiny * MeSH
- uncoupling protein 1 MeSH
The mitochondrial ADP/ATP carrier (AAC, ANT), a member of the SLC25 family of solute carriers, plays a critical role in transporting purine nucleotides (ATP and ADP) as well as protons across the inner mitochondrial membrane. However, the precise mechanism and physiological significance of proton transport by ADP/ATP carrier remain unclear. Notably, the presence of uncouplers-such as long-chain fatty acids (FA) or artificial compounds like dinitrophenol (DNP)-is essential for this process. We explore two potential mechanisms that describe ADP/ATP carrier as either (i) a proton carrier that functions in the presence of FA or DNP, or (ii) an anion transporter (FA- or DNP). In the latter case, the proton is translocated by the neutral form of FA, which carries it from the matrix to the intermembrane space (FA-cycling hypothesis). Our recent results support this hypothesis. We describe a four-step mechanism for the "sliding" of the FA anion from the matrix to the mitochondrial intermembrane space and discuss a possible generalization of this mechanism to other SLC25 carriers.
- Klíčová slova
- MD simulations, bilayer lipid membranes, membrane proteins, mitochondrial transporter, reconstituted protein, uncoupling protein,
- MeSH
- 2,4-dinitrofenol metabolismus MeSH
- adenosintrifosfát metabolismus MeSH
- biologický transport MeSH
- iontový transport MeSH
- lidé MeSH
- mastné kyseliny * metabolismus MeSH
- mitochondriální ADP/ATP-translokasy * metabolismus chemie MeSH
- mitochondrie * metabolismus MeSH
- protony * MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
- Názvy látek
- 2,4-dinitrofenol MeSH
- adenosintrifosfát MeSH
- mastné kyseliny * MeSH
- mitochondriální ADP/ATP-translokasy * MeSH
- protony * MeSH
