NMR characterization and membrane interactions of the loop region of kindlin-3 F1 subdomain

Kindlins-1,2 and 3 are FERM domain-containing cytosolic proteins involved in the activation and regulation of integrin-mediated cell adhesion. Apart from binding to integrin β cytosolic tails, kindlins and the well characterized integrin-activator talin bind membrane phospholipids. The ubiquitin-lik...

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Main Authors: Chua, Geok-Lin, Tan, Suet-Mien, Bhattacharjya, Surajit
Other Authors: Mofrad, Mohammad R. K.
Format: Article
Language:English
Published: 2018
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Online Access:https://hdl.handle.net/10356/80464
http://hdl.handle.net/10220/46562
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Institution: Nanyang Technological University
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spelling sg-ntu-dr.10356-804642023-02-28T16:59:45Z NMR characterization and membrane interactions of the loop region of kindlin-3 F1 subdomain Chua, Geok-Lin Tan, Suet-Mien Bhattacharjya, Surajit Mofrad, Mohammad R. K. School of Biological Sciences DRNTU::Science::Biological sciences Membrane Interactions F1 Loop Kindlins-1,2 and 3 are FERM domain-containing cytosolic proteins involved in the activation and regulation of integrin-mediated cell adhesion. Apart from binding to integrin β cytosolic tails, kindlins and the well characterized integrin-activator talin bind membrane phospholipids. The ubiquitin-like F1 sub-domain of the FERM domain of talin contains a short loop that binds to the lipid membrane. By contrast, the F1 sub-domain of kindlins contains a long loop demonstrated binding to the membrane. Here, we report structural characterization and lipid interactions of the 83-residue F1 loop of kindlin-3 using NMR and optical spectroscopy methods. NMR studies demonstrated that the F1 loop of kindlin-3 is globally unfolded but stretches of residues assuming transient helical conformations could be detected in aqueous solution. We mapped membrane binding interactions of the F1 loop with small unilamellar vesicles (SUVs) containing either zwitterionic lipids or negatively charged lipids using 15N-1H HSQC titrations. These experiments revealed that the F1 loop of kindlin-3 preferentially interacted with the negatively charged SUVs employing almost all of the residues. By contrast, only fewer residues appeared to be interacted with SUVs containing neutral lipids. Further, CD and NMR data suggested stabilization of helical conformations and predominant resonance perturbations of the F1 loop in detergent containing solutions. Conformations of an isolated N-terminal peptide fragment, or EK21, of the F1 loop, containing a poly-Lys sequence motif, important for membrane interactions, were also investigated in detergent solutions. EK21 adopted a rather extended or β-type conformations in complex with negatively charged SDS micelles. To our knowledge, this is the first report describing the conformations and residue-specific interactions of kindlin F1 loop with lipids. These data therefore provide important insights into the interactions of kindlin FERM domain with membrane lipids that contribute toward the integrin activating property. MOE (Min. of Education, S’pore) 2018-11-05T08:11:21Z 2019-12-06T13:50:06Z 2018-11-05T08:11:21Z 2019-12-06T13:50:06Z 2016 Journal Article Chua, G.-L., Tan, S.-M., & Bhattacharjya, S. (2016). NMR characterization and membrane interactions of the loop region of kindlin-3 F1 subdomain. PLOS ONE, 11(4), e0153501-. doi:10.1371/journal.pone.0153501 https://hdl.handle.net/10356/80464 http://hdl.handle.net/10220/46562 10.1371/journal.pone.0153501 27101375 en PLOS ONE © 2016 Chua et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. 14 p. application/pdf
institution Nanyang Technological University
building NTU Library
continent Asia
country Singapore
Singapore
content_provider NTU Library
collection DR-NTU
language English
topic DRNTU::Science::Biological sciences
Membrane Interactions
F1 Loop
spellingShingle DRNTU::Science::Biological sciences
Membrane Interactions
F1 Loop
Chua, Geok-Lin
Tan, Suet-Mien
Bhattacharjya, Surajit
NMR characterization and membrane interactions of the loop region of kindlin-3 F1 subdomain
description Kindlins-1,2 and 3 are FERM domain-containing cytosolic proteins involved in the activation and regulation of integrin-mediated cell adhesion. Apart from binding to integrin β cytosolic tails, kindlins and the well characterized integrin-activator talin bind membrane phospholipids. The ubiquitin-like F1 sub-domain of the FERM domain of talin contains a short loop that binds to the lipid membrane. By contrast, the F1 sub-domain of kindlins contains a long loop demonstrated binding to the membrane. Here, we report structural characterization and lipid interactions of the 83-residue F1 loop of kindlin-3 using NMR and optical spectroscopy methods. NMR studies demonstrated that the F1 loop of kindlin-3 is globally unfolded but stretches of residues assuming transient helical conformations could be detected in aqueous solution. We mapped membrane binding interactions of the F1 loop with small unilamellar vesicles (SUVs) containing either zwitterionic lipids or negatively charged lipids using 15N-1H HSQC titrations. These experiments revealed that the F1 loop of kindlin-3 preferentially interacted with the negatively charged SUVs employing almost all of the residues. By contrast, only fewer residues appeared to be interacted with SUVs containing neutral lipids. Further, CD and NMR data suggested stabilization of helical conformations and predominant resonance perturbations of the F1 loop in detergent containing solutions. Conformations of an isolated N-terminal peptide fragment, or EK21, of the F1 loop, containing a poly-Lys sequence motif, important for membrane interactions, were also investigated in detergent solutions. EK21 adopted a rather extended or β-type conformations in complex with negatively charged SDS micelles. To our knowledge, this is the first report describing the conformations and residue-specific interactions of kindlin F1 loop with lipids. These data therefore provide important insights into the interactions of kindlin FERM domain with membrane lipids that contribute toward the integrin activating property.
author2 Mofrad, Mohammad R. K.
author_facet Mofrad, Mohammad R. K.
Chua, Geok-Lin
Tan, Suet-Mien
Bhattacharjya, Surajit
format Article
author Chua, Geok-Lin
Tan, Suet-Mien
Bhattacharjya, Surajit
author_sort Chua, Geok-Lin
title NMR characterization and membrane interactions of the loop region of kindlin-3 F1 subdomain
title_short NMR characterization and membrane interactions of the loop region of kindlin-3 F1 subdomain
title_full NMR characterization and membrane interactions of the loop region of kindlin-3 F1 subdomain
title_fullStr NMR characterization and membrane interactions of the loop region of kindlin-3 F1 subdomain
title_full_unstemmed NMR characterization and membrane interactions of the loop region of kindlin-3 F1 subdomain
title_sort nmr characterization and membrane interactions of the loop region of kindlin-3 f1 subdomain
publishDate 2018
url https://hdl.handle.net/10356/80464
http://hdl.handle.net/10220/46562
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