Determine both the conformation and orientation of a specific residue in α-synuclein(61–95) even in monolayer by (13)C isotopic label and p-polarized multiple-angle incidence resolution spectrometry (pMAIRS)

Protein’s magic function stems from its structure and various analytical techniques have been developed for it. Among proteins, membrane proteins are encoded 20–30% of genomes, whereas cause challenges for many analytical techniques. For example, lots of membrane proteins cannot form single crystal structure required by X-ray crystallography. As for NMR, the measurements were hindered by the low tumbling rates of membrane (i.e., phospholipid bilayers) where membrane proteins exist. In addition, membrane proteins usually lay parallel to the surface of phospholipid bilayers or form transmembrane structure. No matter parallel or perpendicular to phospholipid bilayers surface, membrane proteins form monolayer structure which is also difficult for X-ray and NMR to provide high-resolution results. Because NMR and X-ray crystallography are the two major analytical techniques to address protein’s structure, membrane proteins only contribute 2.4% to the solved protein databank. Surface FT-IR techniques can evaluate the conformation and orientation of membrane proteins by amide I band. Specifically for α-helical peptides/proteins, the orientation of the axis is critical to decide whether proteins form transmembrane structure. Notice that the traditional FT-IR can only provide “low-resolution” results. Here, (13)C isotope was introduced into the nonamyloid component (NAC), which spans residues 61–95 of α-synuclein (α-syn). Then, p-polarized multiple-angle incidence resolution spectrometry (pMAIRS) was used to determine the orientation of a specific residue of α-helical NAC in monolayer. In general, pMAIRS is a novel technique to work complementary with X-ray and NMR to address membrane peptides/proteins structure with high resolution even in monolayer. GRAPHICAL ABSTRACT: [Image: see text] SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s44211-022-00128-0.