During the catalytic cycle of enzymatic reactions, the recognition of a specific substrate by the enzyme is one of the most important steps, and the analysis of the affinity between enzyme and substrates and/or inhibitors is an important aspect in mechanistic biochemistry. Here, we have performed an analysis of the binding affinity between two different peptides and the bacterial oligosaccharyltransferase, PglB. We have performed MicroScale Themophoresis experiments to determine the Kd values for the interaction between PglB and fluorescently labeled peptides. The results show a high concordance with previously reported values for the same interaction which were determined by fluorescence anisotropy, showing that MST is a suitable tool for the study of membrane protein-substrate interactions in detergent solutions.
Working with tricky membrane proteins?
NanoTemper tools can help
See more related content
Optimization of membrane proteins for cryo-EM: Screening of copolymer nanodiscs with Prometheus Panta
Membrane proteins present challenges for structural analysis by cryo-EM; Screening of synthetic copolymers with nanoDSF and DLS on Prometheus Panta help select the best solubilization conditions.
20:31The power of Monolith X for Characterizing GPCRs: Accelerating Binding Affinity Measurements
To understand signaling pathways, Spectral Shift technology allows for accurate bindings affinity measurements of membrane proteins like GPCRs in close to native conditions.
14:01The desire to use technology that allows you to study membrane proteins in close-to-native conditions is specifically important when it comes to interaction measurements. It’s always a question how mu
12:06What you’ll learn It’s very common to experience low expression levels when producing membrane proteins — plus they’re finicky and unstable when they're outside of their native environment. Just know
4:09Working with difficult targets like GPCRs and other membrane proteins? Because they live in the cell membrane, are hydrophobic and insoluble, have low expression levels, and require particular deter
25:40Are you struggling to tame your mischievous membrane proteins? Find out how NanoTemper can help you to characterize even the most challenging membrane protein samples. You will learn how Prometheus ca
Quantifying the interaction of phosphite with ABC transporters: MicroScale Thermophoresis and a novel his-tag labeling approach
nanoDSF: Label-free thermal unfolding assay of G Protein-coupled receptors for compound screening and buffer composition optimization
Thermophoretic analysis of ligand-specific conformational states of the inhibitory glycine receptor embedded in copolymer nanodiscs
A protein superfamily meets its match in nanoDSF
G protein-coupled receptors (GPCRs) are the largest class of targets for drug discovery, and the fourth largest superfamily in the human genome. They have been linked to the regulation of many...
Phospholipid membrane fluidity alters ligand binding activity of a G protein-coupled receptor by shifting the conformational equilibrium
Phosphorylation of human aquaporin 2 (AQP2) allosterically controls its interaction with the lysosomal trafficking protein LIP5
Microscale thermophoresis quantifies biomolecular interactions under previously challenging conditions
Structure of DtpA from E. coli in complex with Valganciclovir provides insights into drug binding of human PepT1
