Nanoparticles and photochemistry for native-like transmembrane protein footprinting

Abstract

Mass spectrometry-based footprinting can probe higher order structure of soluble proteins in their native states and serve as a complement to high-resolution approaches. Traditional footprinting approaches, however, are hampered for integral membrane proteins because their transmembrane regions are not accessible to solvent, and they contain hydrophobic residues that are generally unreactive with most chemical reagents. To address this limitation, we bond photocatalytic titanium dioxide (TiO2) nanoparticles to a lipid bilayer. Upon laser irradiation, the nanoparticles produce local concentrations of radicals that penetrate the lipid layer, which is made permeable by a simultaneous laser-initiated Paternò–Büchi reaction. This approach achieves footprinting for integral membrane proteins in liposomes, helps locate both ligand-binding residues in a transporter and ligand-induced conformational changes, and reveals structural aspects of proteins at the flexible unbound state. Overall, this approach proves effective in intramembrane footprinting and forges a connection between material science and biology.

A Novel Signaling Strategy for an Ultrasensitive Photoelectrochemical Immunoassay Based on Electro-Fenton Degradation of Liposomes on a Photoelectrode

“…Lip vesicles with an average particle size of 200 nm were prepared (Figure S7) and then coated on the Au@TiO 2 /FTO electrode. Ti–P bonds can form between liposomes and TiO 2 , so Lip vesicles can be tightly attached on the Au@TiO 2 /FTO electrode. Before the coating of Lip vesicles on Au@TiO 2 /FTO, Au@TiO 2 nanoparticles are clearly observed on the FTO (Figure a).…”

Advances in mass spectrometry‐based footprinting of membrane proteins

“…Recently, one strategy named NanoPOMP was designed to meet these challenges and to achieve increased footprinting coverage over the TM regions of two integral MPs [71]. Briefly, NanoPOMP is suitable for MPs reconstituted in liposomes.…”

“…(C) The location of hydroxyl radical (red) generated from activation of water by X-ray irradiation. Reproduced with permission from ref[70] and the International Union of Crystallography(1) Recently, one strategy named NanoPOMP was designed to meet these challenges and to achieve increased footprinting coverage over the TM regions of two integral MPs[71]. Briefly, NanoPOMP is suitable for MPs reconstituted in liposomes.…”

Investigating Daptomycin-Membrane Interactions Using Native MS and Fast Photochemical Oxidation of Peptides in Nanodiscs

“…17 Other rapid footprinting chemistries have also been successfully used for membrane proteins, including the trifluoromethyl radical (•CF3), carbenes derived from diazirines, and carbocations. [18][19][20][21][22][23][24][25] Slower covalent labeling reagents, including glycine ethyl ester 26 diethylpyrocarbonate 27 have also proven effective. The field is rapidly advancing with the development of new reagents that differ in reactivities, kinetics, and selectivity to enhance structural investigations of diverse targets.…”