The interaction affinity between vascular cell adhesion molecule-1 (VCAM-1) and very late antigen-4 (VLA-4) analyzed by quantitative FRET

PloS One
Sandeep ChakrabortyArthur Chiou

Abstract

Very late antigen-4 (VLA-4), a member of integrin superfamily, interacts with its major counter ligand vascular cell adhesion molecule-1 (VCAM-1) and plays an important role in leukocyte adhesion to vascular endothelium and immunological synapse formation. However, irregular expressions of these proteins may also lead to several autoimmune diseases and metastasis cancer. Thus, quantifying the interaction affinity of the VCAM-1/VLA-4 interaction is of fundamental importance in further understanding the nature of this interaction and drug discovery. In this study, we report an 'in solution' steady state organic fluorophore based quantitative fluorescence resonance energy transfer (FRET) assay to quantify this interaction in terms of the dissociation constant (Kd). We have used, in our FRET assay, the Alexa Fluor 488-VLA-4 conjugate as the donor, and Alexa Fluor 546-VCAM-1 as the acceptor. From the FRET signal analysis, Kd of this interaction was determined to be 41.82 ± 2.36 nM. To further confirm our estimation, we have employed surface plasmon resonance (SPR) technique to obtain Kd = 39.60 ± 1.78 nM, which is in good agreement with the result obtained by FRET. This is the first reported work which applies organic fluorophore ba...Continue Reading

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Citations

Jun 25, 2016·Particle and Fibre Toxicology·Cristhiam Rueda-RomeroErnesto Alfaro-Moreno
Oct 11, 2017·The Journal of Immunology : Official Journal of the American Association of Immunologists·Yajing LuRui Zhou
Jun 9, 2020·Frontiers in Oncology·Erica J Hoffmann, Suzanne M Ponik
Jun 3, 2021·International Journal of Molecular Sciences·Veronica RomanoAlessandro Arcucci
Oct 30, 2021·American Journal of Transplantation : Official Journal of the American Society of Transplantation and the American Society of Transplant Surgeons·Danh T TranSatish N Nadig

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Methods Mentioned

BETA
isothermal
surface
FRET
Fluorescence
chip
surface plasmon resonance
chips

Software Mentioned

BIAcore
BIAcore analysis

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