Agrin is a key organizer of acetylcholine receptor (AChR) clustering at the neuromuscular junction. The binding of agrin to laminin is required for its localization to synaptic basal lamina and other basement membranes. The high-affinity interaction with the coiled-coil domain of laminin is mediated by the N-terminal domain of agrin. We have adopted a structurally guided site-directed mutagenesis approach to map the laminin-binding site of NtA. Mutations of L117 and V124 in the C-terminal helix 3 showed that they are crucial for binding. Both residues are located in helix 3 and face the groove between the beta-barrel and the C-terminal helical segment of NtA. Remarkably, the distance between both residues matches a heptad repeat distance of two aliphatic residues which are solvent exposed in the coiled-coil domain of laminin. A lower but significant contribution originates from R43 and a charged cluster (E23, E24 and R40) at the open face of the beta-barrel structure. We propose that surface-exposed, conserved residues of the laminin gamma1 chain interact with NtA via hydrophobic and ionic interactions.
机构:NCI, Frederick Canc Res & Dev Ctr, ABL Basic Res Program, Macromol NMR Sect, Frederick, MD 21702 USA
Zhou, HJ
Mazzulla, MJ
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机构:NCI, Frederick Canc Res & Dev Ctr, ABL Basic Res Program, Macromol NMR Sect, Frederick, MD 21702 USA
Mazzulla, MJ
Kaufman, JD
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机构:NCI, Frederick Canc Res & Dev Ctr, ABL Basic Res Program, Macromol NMR Sect, Frederick, MD 21702 USA
Kaufman, JD
Stahl, SJ
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机构:NCI, Frederick Canc Res & Dev Ctr, ABL Basic Res Program, Macromol NMR Sect, Frederick, MD 21702 USA
Stahl, SJ
Wingfield, PT
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机构:NCI, Frederick Canc Res & Dev Ctr, ABL Basic Res Program, Macromol NMR Sect, Frederick, MD 21702 USA
Wingfield, PT
Rubin, JS
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机构:NCI, Frederick Canc Res & Dev Ctr, ABL Basic Res Program, Macromol NMR Sect, Frederick, MD 21702 USA
Rubin, JS
Bottaro, DP
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机构:NCI, Frederick Canc Res & Dev Ctr, ABL Basic Res Program, Macromol NMR Sect, Frederick, MD 21702 USA
Bottaro, DP
Byrd, RA
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NCI, Frederick Canc Res & Dev Ctr, ABL Basic Res Program, Macromol NMR Sect, Frederick, MD 21702 USANCI, Frederick Canc Res & Dev Ctr, ABL Basic Res Program, Macromol NMR Sect, Frederick, MD 21702 USA
机构:
CEA, DSV, IBEB, Lab Interact Prot Met, St Paul Les Durance, France
CNRS, UMR Biol Vegetale & Microbiol Environm, St Paul Les Durance, France
Aix Marseille Univ, BVME, F-13284 Marseille, FranceCEA, DSV, IBEB, Lab Interact Prot Met, St Paul Les Durance, France
Beccia, Maria Rosa
Sauge-Merle, Sandrine
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CEA, DSV, IBEB, Lab Interact Prot Met, St Paul Les Durance, France
CNRS, UMR Biol Vegetale & Microbiol Environm, St Paul Les Durance, France
Aix Marseille Univ, BVME, F-13284 Marseille, FranceCEA, DSV, IBEB, Lab Interact Prot Met, St Paul Les Durance, France
Sauge-Merle, Sandrine
Lemaire, David
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CEA, DSV, IBEB, Lab Interact Prot Met, St Paul Les Durance, France
CNRS, UMR Biol Vegetale & Microbiol Environm, St Paul Les Durance, France
Aix Marseille Univ, BVME, F-13284 Marseille, FranceCEA, DSV, IBEB, Lab Interact Prot Met, St Paul Les Durance, France
Lemaire, David
Bremond, Nicolas
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CEA, DSV, IBEB, Lab Interact Prot Met, St Paul Les Durance, France
CNRS, UMR Biol Vegetale & Microbiol Environm, St Paul Les Durance, France
Aix Marseille Univ, BVME, F-13284 Marseille, FranceCEA, DSV, IBEB, Lab Interact Prot Met, St Paul Les Durance, France
Bremond, Nicolas
Pardoux, Romain
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CEA, DSV, IBEB, Lab Interact Prot Met, St Paul Les Durance, France
CNRS, UMR Biol Vegetale & Microbiol Environm, St Paul Les Durance, France
Aix Marseille Univ, BVME, F-13284 Marseille, FranceCEA, DSV, IBEB, Lab Interact Prot Met, St Paul Les Durance, France
Pardoux, Romain
Blangy, Stephanie
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Aix Marseille Univ, Architecture & Fonct Macromol Biol, F-13284 Marseille 09, France
CNRS, Architecture & Fonct Macromol Biol UMR7257, F-13288 Marseille 09, FranceCEA, DSV, IBEB, Lab Interact Prot Met, St Paul Les Durance, France
Blangy, Stephanie
Guilbaud, Philippe
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CEA, DRCPC, SMCS, LILA, Bagnols Sur Ceze, FranceCEA, DSV, IBEB, Lab Interact Prot Met, St Paul Les Durance, France
Guilbaud, Philippe
Berthomieu, Catherine
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CEA, DSV, IBEB, Lab Interact Prot Met, St Paul Les Durance, France
CNRS, UMR Biol Vegetale & Microbiol Environm, St Paul Les Durance, France
Aix Marseille Univ, BVME, F-13284 Marseille, FranceCEA, DSV, IBEB, Lab Interact Prot Met, St Paul Les Durance, France