Recombinant yeast VDAC2: a comparison of electrophysiological features with the native form

• Published in: FEBS open bio 2019-07, Vol.9 (7), p.1184-1193
• Main Authors: Magrì, Andrea, Karachitos, Andonis, Di Rosa, Maria Carmela, Reina, Simona, Conti Nibali, Stefano, Messina, Angela, Kmita, Hanna, De Pinto, Vito
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Channel gating; Chromatography; Electrophysiological Phenomena; electrophysiology; Experiments; heterologous expression; Intracellular Membranes - metabolism; Lipid bilayers; Lipid Bilayers - metabolism; Membranes; Mitochondria; Mitochondria - metabolism; mitochondrial porins; planar lipid bilayer; Protein Engineering - methods; Protein expression; Protein Isoforms - metabolism; Proteins; Recombinant Proteins - metabolism; Saccharomyces cerevisiae - metabolism; Saccharomyces cerevisiae Proteins - metabolism; Software; VDAC2; Voltage; Voltage-Dependent Anion Channel 2 - metabolism; Voltage-Dependent Anion Channel 2 - physiology; Voltage-Dependent Anion Channels - metabolism; Yeast; yeastVDAC2; yVDAC2
• ISSN: 2211-5463; 2211-5463
• DOI: 10.1002/2211-5463.12574

Voltage‐dependent anion channel isoform 2 of the yeast Saccharomyces cerevisiae (yVDAC2) was believed for many years to be devoid of channel activity. Recently, we isolated yVDAC2 and showed that it exhibits channel‐forming activity in the planar lipid bilayer system when in its so‐called native form. Here, we describe an alternative strategy for yVDAC2 isolation, through heterologous expression in bacteria and refolding in vitro. Recombinant yVDAC2, like its native form, is able to form voltage‐dependent channels. However, some differences between native and recombinant yVDAC2 emerged in terms of voltage dependence and ion selectivity, suggesting that, in this specific case, the recombinant protein might be depleted of post‐translational modification(s) that occur in eukaryotic cells. The second isoform of yeast mitochondrial porin, yVDAC2, was believed to be devoid of channel activity for many years, up to the recent electrophysiological characterization of the mitochondrial‐extracted protein. Here, a similar pore‐forming activity was demonstrated for the recombinant yVDAC2. At the same time, several features were noticed suggesting the presence of post‐translational modifications occurring in the native protein.