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dc.contributor.advisorPil Seok Chae-
dc.description.abstractthese agents formed PDCs much smaller than DDM. This is a notable observation because both detergent characteristics (small PDC formation and enhanced protein stabilization) are not often compatible with each other within a single molecule. The third novel agents were designed by linking two malonate-based scaffolds. This class could be categorized into two groups: alkyl-based (TMG-As) and thio-ether functionalized TMGs (TMG-Ts). When these agents were evaluated with five membrane proteins including β2AR, we found that some TMGs conferred enhanced stability to all the tested proteins as compared to the gold standard detergent, DDM. This result is marked because it is very challenging to develop novel glucosides with enhanced protein stabilization efficacy toward a range of membrane proteins. As the original MNAs appeared to have long alkyl chains, we have prepared new MNAs with a shorter alkyl chain length by introducing (a) branch point(s) in the alkyl chain with keeping the number of carbons (i.e., hydrophobicity). Various hydrophobic groups including a saturated farnesyl group, symmetric and asymmetric C20 to C24 were conjugated into the hydrophilic group. Effect of detergent hydrophobic density and alkyl chain length on membrane protein stability will be a main topic in this evaluation. As maltoside head group is generally stabilizing protein better than glucoside, we also prepared maltoside versions of the MNAs. Because of the large hydrophilicity, large hydrophobic groups such as branched alkyl chains (C-20, C-22, and C-24) or steroidal unit (e.g., cholesterol) were introduced as detergent tail groups. These glucoside and maltoside variants were compared with the original MNAs for membrane protein stabilization to find detergent structure-property-efficacy relationship.-
dc.description.abstractIntegral membrane proteins (IMPs) are amphipathic macromolecules crucial for a variety of cellular processes. These macromolecules play a central role in materials transport, signal transduction, cell recognition and communication, and many others. The fact that more than 50% pharmaceuticals are designed to target these proteins indicates the importance of these biomolecules in drug discovery. Despite their biological and pharmaceutical importance, less than 1% of these biomolecules are structurally characterized in an atomic resolution. The structural and functional understanding of IMPs is hampered by the difficulty in handling these biomolecules in an aqueous medium. Protein extraction from the membranes, necessary for the downstream characterization, typically needs a detergent to shield the large hydrophobic surfaces of IMPs and as a result to prevent protein denaturation and aggregation. Many membrane proteins remain difficult to study owing to the lack of suitable detergents for their effective solubilization and stabilization. New amphiphilic agents capable of maintaining protein solubility and stability will be helpful to overcome the barrier in membrane protein structural analysis. In the current study we have designed and synthesized novel classes of amphiphiles and their variants targeted for membrane protein structural study. These novel classes are divided into three classes: resorcinarene-based amphiphiles (RGAs/RMAs), mannitol-based amphiphiles (MNAs) and tandem malonate-based glucosides (TMGs). The first novel agents (i.e., resorcinarene-based amphiphiles) contain glucoside (RGAs) or maltoside head group (RMAs). Both sets bear four alkyl chains, facially attached to the central scaffold. However, these two groups differ from each other in the rigidity of the molecules-
dc.description.abstractthe RGAs are more flexible than the RMAs mainly due to the absence of the upper connection around central resorcinarene scaffold. When these agents were tested with four membrane proteins including a G protein-coupled receptor (GPCR), the RGAs overall showed more promising behaviors toward protein stabilization as compared to the RMAs. Of these novel agents, RGA-C11/13 showed the most favorable behaviors compared to DDM. The promising result of the RGAs compared to RMAs indicates that detergent structural flexibility is necessary to effectively stabilize membrane protein structure. The second novel agents prepared for membrane protein study was mannitol-based amphiphiles (MNAs), which were synthesized by connecting four glucose units to the two alkyl chains via a central mannitol linker. The alkyl chain length varied from C8 to C14 to find an optimal hydrophile-liphophile balance, known to be critical for membrane protein stabilization. When these agents were tested with five representatives of membrane proteins including R. capsultus superassembly, LeuT and β2AR, MNA-C13 was overall most effective at maintaining protein stability followed by MNA-12/14 for all the tested proteins, indicating the favorable architecture of MNA amphiphiles. In addition, these agents tend to form small protein-detergent complexes (PDCs) relative to DDM, a valuable detergent characteristic for membrane protein crystallization. A similar behavior was observed for the RGAs-
dc.titleDesign, Synthesis and Evaluation of Multiple Carbohydrate-bearing Amphiphiles for Membrane Protein Study-
dc.contributor.googleauthorHazrat Hussain-
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