Abstract
Ly-6/uPAR or three-finger proteins (TFPs) contain a disulfide-stabilized β-structural core and three protruding loops (fingers). In mammals, TFPs have been found in epithelium and the nervous, endocrine, reproductive, and immune systems. Here, using heteronuclear NMR, we determined the three-dimensional (3D) structure and backbone dynamics of the epithelial secreted protein SLURP-1 and soluble domains of GPI-anchored TFPs from the brain (Lynx2, Lypd6, Lypd6b) acting on nicotinic acetylcholine receptors (nAChRs). Results were compared with the data about human TFPs Lynx1 and SLURP-2 and snake α-neurotoxins WTX and NTII. Two different topologies of the β-structure were revealed: one large antiparallel β-sheet in Lypd6 and Lypd6b, and two β-sheets in other proteins. α-Helical segments were found in the loops I/III of Lynx2, Lypd6, and Lypd6b. Differences in the surface distribution of charged and hydrophobic groups indicated significant differences in a mode of TFPs/nAChR interactions. TFPs showed significant conformational plasticity: the loops were highly mobile at picosecond-nanosecond timescale, while the β-structural regions demonstrated microsecond-millisecond motions. SLURP-1 had the largest plasticity and characterized by the unordered loops II/III and cis-trans isomerization of the Tyr39-Pro40 bond. In conclusion, plasticity could be an important feature of TFPs adapting their structures for optimal interaction with the different conformational states of nAChRs.
Keywords:
15N-relaxation; Ly-6/uPAR; NMR spectroscopy; backbone dynamics; nicotinic acetylcholine receptors; three-finger proteins.
MeSH terms
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Adaptor Proteins, Signal Transducing / chemistry*
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Adaptor Proteins, Signal Transducing / genetics
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Adaptor Proteins, Signal Transducing / metabolism
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Amino Acid Sequence
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Antigens, Ly / chemistry*
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Antigens, Ly / genetics
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Antigens, Ly / metabolism
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Binding Sites
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Cloning, Molecular
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Elapid Venoms / chemistry
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Elapid Venoms / metabolism
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Escherichia coli / genetics
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Escherichia coli / metabolism
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GPI-Linked Proteins / chemistry*
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GPI-Linked Proteins / genetics
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GPI-Linked Proteins / metabolism
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Gene Expression
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Genetic Vectors / chemistry
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Genetic Vectors / metabolism
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Humans
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Hydrophobic and Hydrophilic Interactions
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Models, Molecular
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Neuropeptides / chemistry*
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Neuropeptides / genetics
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Neuropeptides / metabolism
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Nuclear Magnetic Resonance, Biomolecular
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Protein Binding
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Protein Conformation, alpha-Helical
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Protein Conformation, beta-Strand
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Protein Interaction Domains and Motifs
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Protein Isoforms / chemistry
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Protein Isoforms / genetics
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Protein Isoforms / metabolism
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Receptors, Nicotinic / chemistry*
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Receptors, Nicotinic / genetics
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Receptors, Nicotinic / metabolism
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Recombinant Proteins / chemistry
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Recombinant Proteins / genetics
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Recombinant Proteins / metabolism
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Sequence Alignment
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Sequence Homology, Amino Acid
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Urokinase-Type Plasminogen Activator / chemistry*
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Urokinase-Type Plasminogen Activator / genetics
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Urokinase-Type Plasminogen Activator / metabolism
Substances
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Adaptor Proteins, Signal Transducing
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Antigens, Ly
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Elapid Venoms
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GPI-Linked Proteins
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LYNX1 protein, human
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LYPD6 protein, human
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LYPD6B protein, human
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Neuropeptides
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Protein Isoforms
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Receptors, Nicotinic
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Recombinant Proteins
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SLURP1 protein, human
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Weak toxin, Naja
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Urokinase-Type Plasminogen Activator