J Am Chem Soc.2017 Mar;139(8):3072-3081

Conformationally Preorganized Diastereomeric Norbornane-Based Maltosides for Membrane Protein Study: Implications of Detergent Kink for Micellar Properties

Manabendra Das, Yang Du,§ Orquidea Ribeiro, Parameswaran Hariharan, Jonas S. Mortensen,# Dhabaleswar Patra, Georgios Skiniotis, Claus J. Loland,# Lan Guan, Brian K. Kobilka,§ Bernadette Byrne, and Pil Seok Chae†*
Department of Bionanotechnology, Hanyang University, 426-791, Korea
§Department of Molecular and Cellular Physiology, Stanford University, Stanford, CA 94305, USA
Department of Life Sciences, Imperial College London, London, SW7 2AZ, UK
Department of Cell Physiology and Molecular Biophysics, Center for Membrane Protein Research, School of Medicine, Texas Tech University Health Sciences Center, Lubbock, TX 79430, USA
#Department of Neuroscience and Pharmacology, University of Copenhagen, Copenhagen, DK-2200, Denmark
Life Sciences Institute, University of Michigan, Ann Arbor, MI 48109, USA

Corresponding Author *pchae@hanyang.ac.kr



Membrane proteins comprise nearly 30% of most proteomes and facilitate material transport and signal transduction across cell membranes. One major obstacle to membrane protein structure determination is the selection of a detergent suitable for protein stability in aqueous medium. Herein we designed and prepared norbornane-based maltosides (NBMs) with diastereomeric relationship. These diastereomeric NBMs showed different behaviors toward membrane protein stabilization depending on the presence or absence of a kink in the lipophilic region. Specifically, the NBMs with no kink conferred greater stability to most of membrane protein tested here than the NBMs with a kink. Our findings provide a molecular basis for the effect of detergent kinking on micellar properties and associated proteins stability.

Keywords: diastereomeric amphiphiles, membrane proteins, protein stabilization, amphiphile design, protein structure



Two sets of stereo-isomeric detergents with a norbornane linker were prepared and evaluated for their stabilization efficacy with different membrane proteins including a G-protein coupled receptor (GPCR) and a membrane protein complex (GPCR-Gs complex). In Figure 1 the chemical structures of the NBMs (middle-right) and their Newman projections (extreme right) were shown. The NBMs were derived from isomeric endo-/ exo-norbornene-dimethanol via β-selective glycosylation (inset in circle).




The NBMs were evaluated for their stabilization efficacy with different membrane proteins (leucine transporter, MelBst, UapA and the human β2 adrenergic receptor). The marked stabilization of β2AR-Gs complex was attained in X-NBM-C11, which enabled us to clearly visualize the individual domains of the complex via EM analysis. We found that some of these novel agents conferred strikingly greater stability to the target proteins compared to DDM, with the X-isomers mostly better performing than the D-isomers.

Our study not only describes novel maltoside agents with enhanced protein-stabilizing properties, but also suggests that overall detergent geometry (exo or endo) has an important role in determining membrane protein stability. Stereo-isomeric comparison of X- vs D-NBMs strongly indicates that the presence of detergent kink significantly affects the CMC values of detergents as well as its efficacy toward membrane protein stabilization. Based on this study, we propose that detergent packing density in micellar environments should be considered as a critical factor when a novel agent is designed for membrane protein study.



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