Characterization of structures and interactions of beta 2 integrin tail with cytosolic proteins
Lu, Lewis Zhiping
Date of Issue2018
School of Biological Sciences
Integrins are important transmembrane proteins that facilitate many vital processes in humans such as development, immune responses and blood clotting. Extracellular domains of integrin bind to ligands such as the ECM and to other cells while intra-cellularly, cytoplasmic tail domains interact with a wide variety of regulatory adaptor proteins. The regulatory proteins control the conformation and in turn the availability of the extracellular domains for ligand binding. Two of such regulatory proteins are the filamins and migfilin. Filamins are a group of actin crosslinking proteins that negatively regulate the activity of integrin receptors. Filamin associates with the integrin tails through its many individual immunoglobulin (Ig) domains. Filamin A Ig domain 21 (FLNa21) was found to be the primary β integrin tail interactor. Phosphorylation of β integrin tail was found to control the binding of FLNa21. As the phosphorylation site on the β integrin tail was found to be buried deeply within FLNa21 upon binding, we wonder how cellular kinases can get access to that region. Structural as well as NMR dynamics data were obtained using a chimeric β2-FLNa21 protein in order to address that query. Integrin αMβ2 plays multiple important roles in the immune system as facilitators of leukocyte trafficking, in phagocytosis and also in binding to complement products as well. The molecular mechanism for inhibition of integrin activity by FLNa21 binding to integrin tails had been elucidated for the αIIbβ3-FLNa21 system. However, several discrepancies were found in our study using αMβ2 tails and FLNa21. Hence through multiple NMR titration experiments as well as computational dockings, we proposed a model for the interactions between αMβ2 tails and FLNa21. Migfilin is an indirect integrin activator as it competes with β integrin tail for binding to filamin. Migfilin was found to interact with filamin through a stretch of 15 amino acids at its N-terminus. A 13 residues peptide KP13 was designed based on that region. Alanine walk mutational studies were conducted using the FLNa21 and KP13 peptides in order to characterize the critical residues important for binding.