First 3D-Structural Data of Full-Length Guanylyl Cyclase 1 in Rod-Outer-Segment Preparations of Bovine Retina by Cross-Linking/Mass Spectrometry.

The rod-outer-segment guanylyl cyclase 1 (ROS-GC1) is a key transmembrane protein for retinal phototransduction. Mutations of ROS-GC1 correlate with different retinal diseases that often lead to blindness. No structural data are available for ROS-GC1 so far.

Oligomeric state, hydrodynamic properties and target recognition of human Calcium and Integrin Binding protein 2 (CIB2).

Calcium- and Integrin-Binding protein 2 (CIB2) is a small and ubiquitously expressed protein with largely unknown biological function but ascertained role in hearing physiology and disease. Recent studies found that CIB2 binds Ca with moderate affinity and dimerizes under conditions mimicking the physiological ones. Here we provided new lines of evidence on CIB2 oligomeric state and the mechanism of interaction with the α7B integrin target.

Molecular Details of Retinal Guanylyl Cyclase 1/GCAP-2 Interaction.

The rod outer segment guanylyl cyclase 1 (ROS-GC1) is an essential component of photo-transduction in the retina. In the light-induced signal cascade, membrane-bound ROS-GC1 restores cGMP levels in the dark in a calcium-dependent manner. With decreasing calcium concentration in the intracellular compartment, ROS-GC1 is activated via the intracellular site by guanylyl cyclase-activating proteins (GCAP-1/-2).

The First MS-Cleavable, Photo-Thiol-Reactive Cross-Linker for Protein Structural Studies.

Cleavable cross-linkers are gaining increasing importance for chemical cross-linking/mass spectrometry (MS) as they permit a reliable and automated data analysis in structural studies of proteins and protein assemblies. Here, we introduce 1,3-diallylurea (DAU) as the first CID-MS/MS-cleavable, photo-thiol-reactive cross-linker. DAU is a commercially available, inexpensive reagent that efficiently undergoes an anti-Markovnikov hydrothiolation with cysteine residues in the presence of a radical initiator upon UV-A irradiation.

Carboxyl-Photo-Reactive MS-Cleavable Cross-Linkers: Unveiling a Hidden Aspect of Diazirine-Based Reagents.

A major challenge in cross-linking/mass spectrometry (MS) is targeting carboxyl functions in proteins under physiological conditions that do not disturb the protein's conformation. Cross-linking of glutamic acid and aspartic acid residues in proteins will greatly expand the scope of structural mass spectrometry. We discovered that carboxyl-reactive cross-linkers have already been employed for many years in cross-linking/MS studies, yet in a completely different context.

The First Zero-Length Mass Spectrometry-Cleavable Cross-Linker for Protein Structure Analysis.

Combining the properties of a zero-length cross-linker with cleavability by tandem mass spectrometry (MS/MS) poses great advantages for protein structure analysis using the cross-linking/MS approach. These include a reliable, automated data analysis and the possibility to obtain short-distance information of protein 3D-structures. We introduce 1,1'-carbonyldiimidazole (CDI) as an easy-to-use and commercially available, low-cost reagent that ideally fulfils these features.