Intramolecular disulfide bridges in Voltage-Dependent Anion Channel 2 (VDAC2) protein from Rattus norvegicus revealed by high resolution mass spectrometry.

ABSTRACT: Voltage-Dependent Anion Channel isoforms (VDAC1, VDAC2, VDAC3) are relevant component of the outer mitochondrial membrane and play crucial role in cellular processes, in regulation of metabolism and in survival pathways. Recently, we reported the characterization of the oxidation pattern of cysteines in rat and human VDACs and highlighted the key role of these residues in protein function. However, the presence and localization of disulfide bonds in VDACs, a key point for their structural characterization, has so far remained undetermined. Herein we have investigated by nanoUHPLC/High Resolution nanoESI-MS/MS. the position of intramolecular disulfide bonds in the rat VDAC2 (rVDAC2), a protein that contains 11 cysteines. To this purpose, extraction, purification, and enzymatic digestions were carried out at slightly acidic or neutral pH in order to avoid disulfide bond interchange. The presence of six disulfide bridges was unequivocally determined, including a disulfide bridge linking the two adjacent cysteines 4 and 5, a disulfide bridge linking cysteines 9 and 14 and the alternative disulfide bridges between cysteines 48, 77 and 104. A disulfide bond, very resistant to reduction, between cysteines 134 and 139 was also detected. In addition to the previous findings, these results significantly extend the characterization of the oxidation state of cysteines in rVDAC2 and show that it is highly complex and presents unusual features.

INSTRUMENT(S): Orbitrap Fusion

ORGANISM(S): Rattus Rattus (black Rat)

TISSUE(S): Liver

SUBMITTER: Rosaria Saletti  

LAB HEAD: Rosaria Saletti

PROVIDER: PXD044041 | Pride | 2024-06-16

REPOSITORIES: Pride