Unraveling unfolding mechanism of B.subtilis NAD synthase by H/D exchange and native electrospray mass spectrometry

摘要

NAD synthase of Bacillus anthracis is considered promising drug target against anthrax. Most structural information on NAD synthase of Bacillus's genus has been obtained using enzyme from non-pathogenic Bacillus subtilis which shares extensive sequence similarity with NAD synthase of B.antracis. Studying unfolding mechanism of proteins is important for elucidating their structural features, mechanism of action and its regulation. NAD synthase of B.subtilis is a tight dimmer with extensive dimmer interface. Previous results indicated that unfolding of NAD synthase of B.subtilis likely proceeds via a three-state sequential unfolding mechanism, the second step being folded monomer. In this work we sought direct evidence of this using H/D exchange/mass spectrometry and native electrospray mass spectrometry.