[KSCI] Korea Science Citation Index Service

http://dx.doi.org/10.14348/molcells.2021.0016

Biochemical Characterization of the Num1-Mdm36 Complex at the Mitochondria-Plasma Membrane Contact Site

Won, Jongdae (Department of Chemistry, College of Natural Sciences, Seoul National University)
Choi, Yuri (Department of Chemistry, College of Natural Sciences, Seoul National University)
Yun, Yaejin (Department of Chemistry, College of Natural Sciences, Seoul National University)
Lee, Hyung Ho (Department of Chemistry, College of Natural Sciences, Seoul National University)

Abstract

In eukaryotic cells, organelles are distributed and positioned in proximity to each other through molecular tether proteins. Among these, the mitochondria-endoplasmic reticulum cortex anchor (MECA) is a well-known tethering complex in Saccharomyces cerevisiae that tethers mitochondria to the plasma membrane and plays a key role in mitochondrial fission. The main components of MECA are Num1 and Mdm36, and it is known that Mdm36 binds to Num1 to enhance mitochondrial tethering. To better understand the biochemical characteristics of the Num1-Mdm36 complex at the molecular level, we purified the coiled-coil domain of Num1, full-length Mdm36, and Num1-Mdm36 complex and identified the oligomeric state and stoichiometric characteristics of the Num1-Mdm36 complex by chemical crosslinking, size-exclusion chromatography coupled with multi-angle light scattering, and isothermal titration calorimetry. Mdm36 exists as a dimer and interacts preferentially with Num1 with a stoichiometry of 2:2, forming a heterotetrameric complex. Furthermore, we narrowed down the specific binding region of Num1, which is essential for interacting with Mdm36, and showed that their binding affinity is strong enough to tether both mitochondrial and plasma membranes. Our biochemical characterizations suggest a stoichiometric model of the Num1-Mdm36 complex at the mitochondria-plasma membrane contact site in budding yeast.

Keywords

Mdm36; mitochondria; Num1; plasma membrane; tethering;

Citations & Related Records

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