Monolinks Matter: Rational Modification of a Cross-Linker for Improved Flexible Protein Structure Modeling

Chemical cross-linking/mass spectrometry (XL-MS) has emerged as a complementary tool for mapping interaction sites within protein networks as well as gaining moderate-resolution native structurale insight with minimal interference. XL-MS technology mostly relies on chemoselective reactions (crosslinking) between protein residues and a linker. Although complementary to crosslinks, monolinks are less commonly recognised as adjuvants for protein structure elucidation. Herein we report the rational design and synthesis of DSSO-carbamate, a novel cross-linker with increased selectivity for monolinks. Monolinks represent useful information for the elucidation of protein conformational changes and recent work has exploited their application in experimental crosslinking MS integrated protein structure prediction algorithms such as XL-MS-AlphaFold2 . its application supported by AlphaFold2 . Herein we report the rational design and synthesis of DSSO-carbamate, a novel cross-linker with increased selectivity for monolinks. Further, we demonstrate DSSO-carbamate’s capacity to facilitate improved protein structure prediction on account of its superior monolink generation.