Longitudinal separation by transverse diffusion in laminar pipe flow (LSTDLPF): an accurate approach for finding equilibrium constants of protein-small molecule binding
Séminaire Chimie IBMM
Dr. Sergey N.
Le Jeudi 20 Septembre 2018 à 16h
ENSCM, Amphithéâtre Mousseron B (campus Balard, 240 av. Émile-Jeanbrau)
Date de début : 2018-09-20 16:00:00
Date de fin : 2018-09-20 17:30:00
Lieu : ENSCM amphi Mousseron B (campus Balard, 240 av E. Jeanbrau)
Intervenant : Dr. Sergey N.
Department of Chemistry and Centre for Research on Biomolecular Interactions, York University, Toronto, Canada
We developed an accurate approach for determining the equilibrium dissociation constant, Kd, of protein-small molecule complexes. Conceptually, in this approach, a short plug of the pre-equilibrated mixture of a protein and a small molecule is propagated by pressure in a laminar pipe flow. Differential transverse diffusion of the protein-small molecule complex and the unbound small molecule leads to their fast transient separation in the longitudinal direction. A signal proportional to the total concentration of the small molecule (protein-bound and unbound) is measured as a function of time at the pipe exit for varying concentrations of the protein and a constant concentration of the small molecule. A classical binding isotherm — fraction of unbound small molecule as a function of protein concentration — is built and used to find Kd. In this lecture, I will explain the fundamentals of our approach as well as demonstrate its experimental proof-in-principle.
Contact local IBMM : Prof. Hervé
