Maltose-bis(hydroxymethyl)phenol (MBPs) and Maltose-tris(hydroxymethyl)phenol (MTPs) Amphiphiles for Membrane Protein Stability

Abstract

Membrane protein structures provide a fundamental understanding of their molecular actions and are of importance for drug development. Detergents are widely used to solubilize, stabilize, and crystallize membrane proteins, but membrane proteins solubilized in conventional detergents are prone to denaturation and aggregation. Thus, developing novel detergents with enhanced efficacy for protein stabilization remains important. We report herein the design and synthesis of a class of phenol-derived maltoside detergents. Using two different linkers, we prepared two sets of new detergents, designated maltose-bis­(hydroxymethyl)­phenol (MBPs) and maltose-tris­(hydroxymethyl)­phenol (MTPs). The evaluation of these detergents with three transporters and two G-protein coupled receptors allowed us to identify a couple of new detergents (MBP-C9 and MTP-C12) that consistently conferred enhanced stability to all tested proteins compared to a gold standard detergent (DDM). Furthermore, the data analysis based on the detergent structures provides key detergent features responsible for membrane protein stabilization that together will facilitate the future design of novel detergents.

Foldable Detergents for Membrane Protein Stability

“…Detergent linkers also dictate the relative directions of head and tail groups, thus affecting detergent geometry and micelle size [20] . Detergent linkers that have been used for this purpose are typically small and branched, as exemplified by the neopentyl glycol (NG), mannitol (MN), scyllo‐inositol (SI), butane‐1,2,3,4‐tetraol (BT), norbornane (NB), and cyclopentane (CP), 1,3,5‐triazine, bis(hydroxymethyl)phenol (BP), pentane‐1,3,5‐triol (PT) linkers used for the neopentyl glycol‐derived triglucosides (NDTs), [21] mannitol‐based amphiphiles (MNAs), [22] scyllo‐inositol‐based trimaltosides (STMs), [23] butane‐1,2,3,4‐tetraol‐based maltosides (BTMs), [24] norbornane‐based maltosides (NBMs), [25] cyclopentane‐based maltosides (CPMs), [26] 1,3,5‐triazine‐based maltosides (TEMs), [27] maltose bis(hydroxymethyl)phenol amphiphiles (MBPs) [28] and 1,3‐acetonedicaboxylate‐derived glucoside amphiphiles (ACAs), [29] respectively (Figure 2). The small and branched linkers play an important role in positioning two or three alkyl chains in close proximity, resulting in a decrease in the inter‐alkyl chain distance and an increase in alkyl chain density inside detergent micelles.…”

Foldable Detergents for Membrane Protein Study: Importance of Detergent Core Flexibility in Protein Stabilization

“…As indicated by the full retention in the substrate binding ability of LeuT during the 13‐day incubation, most TZMs are among the best of the previously reported detergents at stabilising this protein. Specifically, TZM−E9 and TZM−H9 appeared more effective than LMNG, MTG−A13, GNG‐3,14, MBP−C9 and TEM−T10 at stabilizing LeuT long term [29,54,55] . As for MOR stability, the efficacies of TZM−H9 and TZM−E9 appear to be also superior to those of several reported detergents including LMNG, GNG‐3,14, MBP−C9 and TEM−T10; these two TZMs displayed nearly full retention (∼80 %) of the ligand binding ability of the receptor during the 3‐day incubation, while the previous detergents resulted in substantial decreases (<40 %) [29,54,55] .…”

“…Specifically, TZMÀ E9 and TZMÀ H9 appeared more effective than LMNG, MTGÀ A13, GNG-3,14, MBPÀ C9 and TEMÀ T10 at stabilizing LeuT long term. [29,54,55] As for MOR stability, the efficacies of TZMÀ H9 and TZMÀ E9 appear to be also superior to those of several reported detergents including LMNG, GNG-3,14, MBPÀ C9 and TEMÀ T10; these two TZMs displayed nearly full retention (~80 %) of the ligand binding ability of the receptor during the 3-day incubation, while the previous detergents resulted in substantial decreases (< 40 %). [29,54,55] When it comes to β 2 AR stability, the TZMs such as TZMÀ E9 and TZMÀ H9 showed a gradual decrease in ligand binding ability, comparable to the previously reported detergents mentioned above.…”

“…[29,54,55] As for MOR stability, the efficacies of TZMÀ H9 and TZMÀ E9 appear to be also superior to those of several reported detergents including LMNG, GNG-3,14, MBPÀ C9 and TEMÀ T10; these two TZMs displayed nearly full retention (~80 %) of the ligand binding ability of the receptor during the 3-day incubation, while the previous detergents resulted in substantial decreases (< 40 %). [29,54,55] When it comes to β 2 AR stability, the TZMs such as TZMÀ E9 and TZMÀ H9 showed a gradual decrease in ligand binding ability, comparable to the previously reported detergents mentioned above. Taken together, the results obtained for TZMÀ H9 and TZMÀ E9 indicate that these molecules confer greater membrane protein stability compared to DDM and previously developed detergents.…”

Transition‐Linker Containing Detergents for Membrane Protein Studies

“…[10][11] Tremendous efforts have been made to improve detergent efficacy in the stabilization of MPs. Dimerization or oligomerization of traditional detergents, [12][13][14] typically neopentyl- based [15][16] , norbornane-based, [17] cholate-based, [18] triazolebased, [19] and polymeric detergents, [20] seems to reduce the dynamics of micelles and therefore to enhance the stabilization of MPs. Additionally, a class of detergents were also designed to stabilize MPs by optimization of polar groups to clamp the membrane domain, like glycosyl-substituted dicarboxylate detergents.…”