Faculty Collaboration Database

Role of nuclear-encoded subunit Vb in the assembly and stability of cytochrome c oxidase complex: implications in mitochondrial dysfunction and ROS production. Biochem J 2009 May 27;420(3):439-49

Date

04/03/2009

Pubmed ID

19338496

Pubmed Central ID

PMC2735414

DOI

10.1042/BJ20090214

Scopus ID

2-s2.0-66949172391 (requires institutional sign-in at Scopus site)   77 Citations

Abstract

CcO (cytochrome c oxidase) is a multisubunit bigenomic protein complex which catalyses the last step of the mitochondrial electron transport chain. The nuclear-encoded subunits are thought to have roles either in regulation or in the structural stability of the enzyme. Subunit Vb is a peripheral nuclear-encoded subunit of mammalian CcO that is dramatically reduced under hypoxia. Although it has been shown to contain different ligand-binding sites and undergo modifications, its precise function is not known. In the present study we generated a cell line from RAW 264.7 murine macrophages that has a more than 80% reduced level of Vb. Functional analysis of these cells showed a loss of CcO activity, membrane potential and less ability to generate ATP. Resolution of complexes on blue native gel and two-dimensional electrophoretic analysis showed an accumulation of subcomplexes of CcO and also reduced association with supercomplexes of the electron transfer chain. Furthermore, the mitochondria from CcO Vb knock-down cells generated increased ROS (reactive oxygen species), and the cells were unable to grow on galactose-containing medium. Pulse-chase experiments suggest the role of the CcO Vb subunit in the assembly of the complex. We show for the first time the role of a peripheral, non-transmembrane subunit in the formation as well as function of the terminal CcO complex.

Author List

Galati D, Srinivasan S, Raza H, Prabu SK, Hardy M, Chandran K, Lopez M, Kalyanaraman B, Avadhani NG

Author

Balaraman Kalyanaraman PhD Professor in the Biophysics department at Medical College of Wisconsin

MESH terms used to index this publication - Major topics in bold

Adenosine Triphosphate
Animals
Cell Line
Cell Nucleus
Cell Proliferation
Electron Spin Resonance Spectroscopy
Electron Transport
Electron Transport Complex IV
Electrophoresis, Gel, Two-Dimensional
Enzyme Stability
Galactose
Immunoblotting
Macrophages
Membrane Potential, Mitochondrial
Mice
Mitochondria
Oxygen Consumption
Protein Subunits
RNA, Small Interfering
Reactive Oxygen Species
Transfection