NMR and Patch-Clamp Characterization of Yeast Mitochondrial Pyruvate Carrier Complexes
Zhen Wang,
Wen Ding,
Maosen Ruan,
Yong Liu,
Jing Yang,
Huiqin Zhang,
Bing Shen,
Junfeng Wang,
Yunyan Li
Affiliations
Zhen Wang
High Magnetic Field Laboratory, CAS Key Laboratory of High Magnetic Field and Ion Beam Physical Biology, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei 230031, China
Wen Ding
School of Basic Medical Sciences, Anhui Medical University, Hefei 230032, China
Maosen Ruan
High Magnetic Field Laboratory, CAS Key Laboratory of High Magnetic Field and Ion Beam Physical Biology, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei 230031, China
Yong Liu
High Magnetic Field Laboratory, CAS Key Laboratory of High Magnetic Field and Ion Beam Physical Biology, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei 230031, China
Jing Yang
High Magnetic Field Laboratory, CAS Key Laboratory of High Magnetic Field and Ion Beam Physical Biology, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei 230031, China
Huiqin Zhang
High Magnetic Field Laboratory, CAS Key Laboratory of High Magnetic Field and Ion Beam Physical Biology, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei 230031, China
Bing Shen
School of Basic Medical Sciences, Anhui Medical University, Hefei 230032, China
Junfeng Wang
High Magnetic Field Laboratory, CAS Key Laboratory of High Magnetic Field and Ion Beam Physical Biology, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei 230031, China
Yunyan Li
High Magnetic Field Laboratory, CAS Key Laboratory of High Magnetic Field and Ion Beam Physical Biology, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei 230031, China
The mitochondrial pyruvate carrier (Mpc) plays an indispensable role in the transport of pyruvates across the mitochondrial inner membrane. Despite the two distinct homologous proteins, Mpc1 and Mpc2, were identified in 2012, there are still controversies on the basic functional units and oligomeric state of Mpc complexes. In this study, yeast Mpc1 and Mpc2 proteins were expressed in a prokaryotic heterologous system. Both homo- and hetero-dimers were successfully reconstituted in mixed detergents. Interactions among Mpc monomers were recorded utilizing paramagnetic relaxation enhancement (PRE) nuclear magnetic resonance (NMR) methods. By single-channel patch-clamp assays, we discovered that both the Mpc1–Mpc2 hetero-dimer and Mpc1 homo-dimer are able to transport K+ ions. Furthermore, the Mpc1–Mpc2 hetero-dimer demonstrated the ability to transport pyruvates, at a rate significantly higher than that of the Mpc1 homo-dimer, indicating that it could be the basic functional unit of Mpc complexes. Our findings provide valuable insights for further structural determination and the study of the transport mechanism of Mpc complexes.
