1. LRIG1 Extracellular Domain: Structure and Function Analysis
- Author
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Thomas P. J. Garrett, Nicos A. Nicola, Priscilla Soo, Yibin Xu, Francesca Walker, Chin Wee Tan, Timothy E. Adams, Nicholas T. Redpath, Antony W. Burgess, Hui Hua Zhang, and Jian-Guo Zhang
- Subjects
Models, Molecular ,Protein Conformation ,chemical and pharmacologic phenomena ,Biosensing Techniques ,Leucine-rich repeat ,Biology ,Crystallography, X-Ray ,Leucine-Rich Repeat Proteins ,Ligands ,stem cell marker ,Homology (biology) ,Structure-Activity Relationship ,03 medical and health sciences ,0302 clinical medicine ,Protein structure ,Structural Biology ,leucine-rich repeat domain ,Tumor Cells, Cultured ,Extracellular ,Humans ,Structure–activity relationship ,EGFR inhibition ,Molecular Biology ,030304 developmental biology ,0303 health sciences ,Membrane Glycoproteins ,fungi ,HEK 293 cells ,Proteins ,Surface Plasmon Resonance ,Extracellular Matrix ,Protein Structure, Tertiary ,3. Good health ,ErbB Receptors ,HEK293 Cells ,Microscopy, Fluorescence ,Ectodomain ,Biochemistry ,030220 oncology & carcinogenesis ,Colonic Neoplasms ,Biophysics ,LINGO-1 ,Binding domain - Abstract
We have expressed and purified three soluble fragments of the human LRIG1-ECD (extracellular domain): the LRIG1-LRR (leucine-rich repeat) domain, the LRIG1-3Ig (immunoglobulin-like) domain, and the LRIG1-LRR-1Ig fragment using baculovirus vectors in insect cells. The two LRIG1 domains crystallised so that we have been able to determine the three-dimensional structures at 2.3Å resolution. We developed a three-dimensional structure for the LRIG1-ECD using homology modelling based on the LINGO-1 structure. The LRIG1-LRR domain and the LRIG1-LRR-1Ig fragment are monomers in solution, whereas the LRIG1-3Ig domain appears to be dimeric. We could not detect any binding of the LRIG1 domains or the LRIG1-LRR-1Ig fragment to the EGF receptor (EGFR), either in solution using biosensor analysis or when the EGFR was expressed on the cell surface. The FLAG-tagged LRIG1-LRR-1Ig fragment binds weakly to colon cancer cells regardless of the presence of EGFRs. Similarly, neither the soluble LRIG1-LRR nor the LRIG1-3Ig domains nor the full-length LRIG1 co-expressed in HEK293 cells inhibited ligand-stimulated activation of cell-surface EGFR.
- Published
- 2015