title: Modulation of human mitochondrial voltage-dependent anion channel 2 (hVDAC-2) structural stability by cysteine-assisted barrel-lipid interactions.
abstract: Human mitochondrial voltage-dependent anion channel 2 (hVDAC-2), the most predominant isoform seen in brain mitochondria, is not only crucial for cell survival but is also implicated in Alzheimer disease. The abundance of cysteines in this isoform is particularly fascinating, as hVDAC-1 cysteines have no associated functional role. We report a detailed biophysical examination of a Cys-less mutant of hVDAC-2, and its behavioral comparison with the wild type protein. Our findings suggest that cysteine mutation results in the formation of a better barrel at the expense of weakened protein-lipid interactions. The wild type protein displays stronger lipid association, despite being less structured. A reversal in behavior of both proteins is observed in the case of chemical denaturation, with the Cys-less mutant exhibiting lowered unfolding free energies. In bicellar systems comprising 14-C phosphocholines, we observe that protein-lipid interactions are weakened in both constructs, resulting in barrel structure destabilization. Our biochemical and biophysical studies together reveal key structural roles for the cysteine residues. We find that minor conformational variations in local residues are sufficient to define the membrane protein dynamics in hVDAC-2. Such subtle sequence variations contribute to differential stability of VDACs and may have implications in their in vivo regulation and recycling.
authors.affiliation: From the Molecular Biophysics Laboratory, Department of Biological Sciences, Indian Institute of Science Education and Research, Bhopal 462023, India.
authors.forename: Svetlana Rajkumar
authors.initials: SR
authors.lastname: Maurya
authors.affiliation: From the Molecular Biophysics Laboratory, Department of Biological Sciences, Indian Institute of Science Education and Research, Bhopal 462023, India. Electronic address: maha@iiserb.ac.in.
authors.forename: Radhakrishnan
authors.initials: R
authors.lastname: Mahalakshmi
mesh_terms: D000544:Alzheimer Disease / Q000235:genetics / Q000378:metabolism; D003545:Cysteine / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D009154:Mutation; D010767:Phosphorylcholine / Q000737:chemistry* / Q000378:metabolism; D017510:Protein Folding; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D050996:Voltage-Dependent Anion Channel 2 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: C494765:VDAC2 protein, human; D050996:Voltage-Dependent Anion Channel 2; D010767:Phosphorylcholine; D003545:Cysteine
keywords: Anion Channel; Cysteine Mutation; Kinetics; Membrane Biophysics; Membrane Proteins; Protein Denaturation; Protein Folding; Protein-Lipid Interaction; Spectroscopy; Thermodynamics