Optimal Bicelle Size q for Solution NMR Studies of the Protein Transmembrane Partition

Abstract

Structural characterization of transmembrane proteins in isotropic bicelles has become an increasingly popular application of solution NMR spectroscopy, as the fast-tumbling bicelles are membrane-like yet can often yield spectral quality comparable to those of detergent micelles. While larger bicelles are closer to the true lipid bilayer, it remains unclear how large the bicelles need to be to allow accurate assessment of protein transmembrane partition in lipid bilayer. Here, we address the above question from the perspective of protein residing in the bicelles, through systematic measurement of protein chemical shift and transmembrane partition at different lipid:detergent ratios (q), ranging from 0.3 to 0.7, using the transmembrane domain of human Fas receptor as model system. We found that the lipid environment of the bicelles, as reflected by the protein chemical shift, begins to be perturbed when the q is reduced to below 0.6. We also implemented a solvent paramagnetic relaxation enhancement (PRE) approach for bicelles to show that the protein transmembrane partition in bicelles with q = 0.5 and 0.7 are very similar, but at q = 0.3 the solvent PRE profile is significantly different. Our data indicate that q values between 0.5 and 0.6 are good compromise between high resolution NMR and closeness to the membrane environment, and allow accurate characterization of protein position in lipid bilayer.

Structure of the membrane proximal external region of HIV-1 envelope glycoprotein

“…To position the MPER with respect to the lipid bilayer, we analyzed the transmembrane partition of the MPER-TMD trimer using our previously developed paramagnetic probe titration (PPT) method (38). We reconstituted the MPER-TMD protein in bicelles with q = 0.6 and titrated the water-soluble paramagnetic probe Gd-DOTA into the aqueous phase surrounding the bicelles, allowing us to measure residue-specific PRE amplitudes (PRE amp ) ( Fig.…”

“…NMR Structure Determination. We used the following general approach: (i) determine local structure of an individual chain in the oligomer, (ii) apply intermonomer distance restraints to derive the oligomer structure, (iii) crossvalidate the NOE-derived structure using PRE from spin labels, and (iv) determine the transmembrane partition in bicelles by PPT (38). We determined secondary structures of the subunits in the oligomeric complex using local NOE restraints and backbone dihedral restraints derived from chemical shifts and TALOS+ (69).…”

Beyond detergent micelles: The advantages and applications of non-micellar and lipid-based membrane mimetics for solution-state NMR

“…Since their size can be fine-tuned in an easy manner, this system is suitable for a wide range of MPs and has been extensively used for solution NMR studies [28][29][30][31]. More recently, an optimal q-value of 0.5 was determined that leads to the most realistic bilayer properties for solution-state NMR applications [32].…”

“…The benefit of bicelles for solution-state NMR is their tunable size [24]. In a recent study, it was concluded that bicelles with a q-value of 0.5 or above can be considered to be a realistic membrane mimic [32].…”

Solution NMR Studies of Anesthetic Interactions with Ion Channels

“…Although a native membrane environment is the most ideal choice, the large size of lipid vesicles is problematic for solution NMR. Alternative choices include detergent micelles, phospholipid bicelles, and nanodiscs (Piai, Fu, Dev, & Chou, 2017; Sanders & Mittendorf, 2011) (Etzkorn et al, 2013; Hagn, Etzkorn, Raschle, & Wagner, 2013; Kucharska, Edrington, Liang, & Tamm, 2015; Laguerre et al, 2016){Susac, 2014 #693}(Liang & Tamm, 2016). Detergent micelles are often used for solution NMR due to their small size.…”