Subunit NDUFV3 is present in two distinct isoforms in mammalian complex I

Area of Science:

• Mitochondrial biochemistry
• Enzyme complex structure and function
• Proteomics

Background:

• Complex I (NADH:ubiquinone oxidoreductase) is crucial for mitochondrial respiration.
• Previous studies on bovine heart Complex I revealed 45 subunits.
• The existence of additional subunits or isoforms in other species/tissues remained unexplored.

Purpose of the Study:

• To characterize Complex I from various rat tissues and a hepatoma cell line.
• To identify potential novel subunits or isoforms of Complex I.
• To investigate the distribution and stoichiometry of identified isoforms.

Main Methods:

• Purification of Complex I from five rat tissues and a rat hepatoma cell line.
• Liquid chromatography-mass spectrometry (LC-MS) and MALDI-TOF mass spectrometry.
• Blue native polyacrylamide gel electrophoresis (BN-PAGE).

Main Results:

• Identification of a ~50kDa isoform of the NDUFV3 subunit, distinct from the canonical ~10kDa form.
• The long NDUFV3 isoform was detected in mature Complex I from rat tissues and human cells, albeit at substoichiometric levels.
• Higher abundance of the long NDUFV3 isoform was observed in rat brain mitochondria and cultured cells compared to other tissues.

Conclusions:

• A novel, long isoform of NDUFV3 exists in mammalian Complex I.
• This long isoform is present in mature Complex I across species and tissues, with varying abundance.
• The long NDUFV3 isoform may compete with the canonical isoform for binding to the flavoprotein domain of Complex I.