NMR Solution Conformations and Interactions of Integrin αLβ2 Cytoplasmic Tails

Anirban Bhunia,Xiao-Yan Tang,Harini Mohanram,Suet-Mien Tan,Surajit Bhattacharjya

doi:10.1074/jbc.m807236200

Anirban Bhunia, Xiao-Yan Tang + Show 3 more

Open Access

https://doi.org/10.1074/jbc.m807236200

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Abstract

The integrins are bi-directional signal transducers. Devoid of enzymatic activity, the integrin cytoplasmic tail serves as a hub for the recruitment of cytosolic proteins, and many of these are signaling molecules. The leukocyte-restricted integrin alphaLbeta2 is essential for the adhesion, migration, and proliferation of leukocytes. Here we report solution conformations and interactions of the alphaLbeta2 cytoplasmic tails by NMR analyses. The alphaL tail is characterized by three helical segments in the order of helix 1-3 that are connected by two loops with helix 3 having a number of nuclear Overhauser effect contacts with helix 1 and helix 2. The conformation of the beta2 tail is less defined with only a helical segment restricted at its N terminus. Acidic residues from the helix 2-loop-helix 3 motif of alphaL were found to be responsible for its binding to calcium ion. There were detectable interactions between alphaL and beta2 tails, involving helix 1 and helix 3 of the alphaL tail and the N-terminal helix of the beta2 tail. Talin head domain that contains the FERM domain showed binding affinity of Kd approximately 0.5 microm with the beta2 tail. The binding affinity of alphaL and beta2 tails is Kd approximately 2.63 microm. These data are in line with the activating property of talin head domain on alphaLbeta2 by which binding of talin head domain to beta2 tail disrupts the interface of the alphaL and beta2 tails that constrains alphaLbeta2 in a resting state.

Highlights

Resulting in a 1:1.5 molar ratio. 1H-15N heteronuclear single quantum coherence (HSQC) spectra were Resonance Assignments of the Cytoplasmic Tails of ␣L␤2 obtained after addition of talin head domain (0.5 mM) into a Integrin—The ␣L and ␤2 tails were overexpressed into inclusolution of 15N-labeled ␣L tail and unlabeled ␤2 tail
Ments were made in 10 mM phosphate buffer containing 150 mM NaCl, 2 mM TCEP, and pH 6.2 at 293 K
Resulting in a 1:1.5 molar ratio. 1H-15N HSQC spectra were Resonance Assignments of the Cytoplasmic Tails of ␣L␤2 obtained after addition of talin head domain (0.5 mM) into a Integrin—The ␣L and ␤2 tails were overexpressed into inclusolution of 15N-labeled ␣L tail and unlabeled ␤2 tail

Summary

Introduction

Resulting in a 1:1.5 molar ratio. 1H-15N HSQC spectra were Resonance Assignments of the Cytoplasmic Tails of ␣L␤2 obtained after addition of talin head domain (0.5 mM) into a Integrin—The ␣L and ␤2 tails were overexpressed into inclusolution of 15N-labeled ␣L tail and unlabeled ␤2 tail. NOE interactions observed among these residues suggest probable occurrence of such ionic By contrast, the analyses of 13C␣ chemical shift that deviates and/or hydrogen bond formations in ␣L structure (Table 1).

Results

Conclusion