TY  - JOUR
A1  - Schulte, Clemens
A1  - Soldà, Alice
A1  - Spänig, Sebastian
A1  - Adams, Nathan
A1  - Bekić, Ivana
A1  - Streicher, Werner
A1  - Heider, Dominik
A1  - Strasser, Ralf
A1  - Maric, Hans Michael
T1  - Multivalent binding kinetics resolved by fluorescence proximity sensing
JF  - Communications Biology
N2  - Multivalent protein interactors are an attractive modality for probing protein function and exploring novel pharmaceutical strategies. The throughput and precision of state-of-the-art methodologies and workflows for the effective development of multivalent binders is currently limited by surface immobilization, fluorescent labelling and sample consumption. Using the gephyrin protein, the master regulator of the inhibitory synapse, as benchmark, we exemplify the application of Fluorescence proximity sensing (FPS) for the systematic kinetic and thermodynamic optimization of multivalent peptide architectures. High throughput synthesis of +100 peptides with varying combinatorial dimeric, tetrameric, and octameric architectures combined with direct FPS measurements resolved on-rates, off-rates, and dissociation constants with high accuracy and low sample consumption compared to three complementary technologies. The dataset and its machine learning-based analysis deciphered the relationship of specific architectural features and binding kinetics and thereby identified binders with unprecedented protein inhibition capacity; thus, highlighting the value of FPS for the rational engineering of multivalent inhibitors.
KW  - combinatorial libraries
KW  - kinetics
KW  - peptides
KW  - screening
KW  - thermodynamics
Y1  - 2022
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-301157
VL  - 5
ER  -