title: Sequential pH-driven dimerization and stabilization of the N-terminal domain enables rapid spider silk formation.
abstract: The mechanisms controlling the conversion of spider silk proteins into insoluble fibres, which happens in a fraction of a second and in a defined region of the silk glands, are still unresolved. The N-terminal domain changes conformation and forms a homodimer when pH is lowered from 7 to 6; however, the molecular details still remain to be determined. Here we investigate site-directed mutants of the N-terminal domain from Euprosthenops australis major ampullate spidroin 1 and find that the charged residues D40, R60 and K65 mediate intersubunit electrostatic interactions. Protonation of E79 and E119 is required for structural conversions of the subunits into a dimer conformation, and subsequent protonation of E84 around pH 5.7 leads to the formation of a fully stable dimer. These residues are highly conserved, indicating that the now proposed three-step mechanism prevents premature aggregation of spidroins and enables fast formation of spider silk fibres in general.
authors: KI Alzheimer Disease Research Centre, NVS Department, Karolinska Institutet, Novum, 5th floor, SE-141 86 Stockholm, Sweden.
authors: Nina
authors: N
authors: Kronqvist
authors: Department of Physical Organic Chemistry, Latvian Institute of Organic Synthesis, Aizkraukles 21, Riga LV-1006, Latvia.
authors: Martins
authors: M
authors: Otikovs
authors: Experimental Biomolecular Physics, Royal Institute of Technology-KTH, Albanova, SE-106 91 Stockholm, Sweden.
authors: Volodymyr
authors: V
authors: Chmyrov
authors: Institute of Biological Sciences and Biotechnology, Donghua University, Shanghai CN-201620, China.
authors: Gefei
authors: G
authors: Chen
authors: Department of Anatomy, Physiology and Biochemistry, Swedish University of Agricultural Sciences, The Biomedical Centre, SE-751 23 Uppsala, Sweden.
authors: Marlene
authors: M
authors: Andersson
authors: KI Alzheimer Disease Research Centre, NVS Department, Karolinska Institutet, Novum, 5th floor, SE-141 86 Stockholm, Sweden.
authors: Kerstin
authors: K
authors: Nordling
authors: Department of Medical Biochemistry and Biophysics, Karolinska Institutet, SE-171 77 Stockholm, Sweden.
authors: Michael
authors: M
authors: Landreh
authors: KI Alzheimer Disease Research Centre, NVS Department, Karolinska Institutet, Novum, 5th floor, SE-141 86 Stockholm, Sweden.
authors: Médoune
authors: M
authors: Sarr
authors: Department of Medical Biochemistry and Biophysics, Karolinska Institutet, SE-171 77 Stockholm, Sweden.
authors: Hans
authors: H
authors: Jörnvall
authors: 1] Experimental Biomolecular Physics, Royal Institute of Technology-KTH, Albanova, SE-106 91 Stockholm, Sweden [2] Science for Life Laboratory, SE-171 65 Solna, Sweden.
authors: Stefan
authors: S
authors: Wennmalm
authors: Experimental Biomolecular Physics, Royal Institute of Technology-KTH, Albanova, SE-106 91 Stockholm, Sweden.
authors: Jerker
authors: J
authors: Widengren
authors: Institute of Biological Sciences and Biotechnology, Donghua University, Shanghai CN-201620, China.
authors: Qing
authors: Q
authors: Meng
authors: 1] KI Alzheimer Disease Research Centre, NVS Department, Karolinska Institutet, Novum, 5th floor, SE-141 86 Stockholm, Sweden [2] Department of Anatomy, Physiology and Biochemistry, Swedish University of Agricultural Sciences, The Biomedical Centre, SE-751 23 Uppsala, Sweden.
authors: Anna
authors: A
authors: Rising
authors: Center for Insoluble Protein Structures (inSPIN), iNANO and Department of Molecular Biology and Genetics, Aarhus University, Gustav Wieds Vej 14, DK-8000 Aarhus C, Denmark.
authors: Daniel
authors: D
authors: Otzen
authors: Department of Cell and Molecular Biology, Uppsala University, SE-751 24 Uppsala, Sweden.
authors: Stefan D
authors: SD
authors: Knight
authors: Department of Physical Organic Chemistry, Latvian Institute of Organic Synthesis, Aizkraukles 21, Riga LV-1006, Latvia.
authors: Kristaps
authors: K
authors: Jaudzems
authors: 1] KI Alzheimer Disease Research Centre, NVS Department, Karolinska Institutet, Novum, 5th floor, SE-141 86 Stockholm, Sweden [2] Department of Anatomy, Physiology and Biochemistry, Swedish University of Agricultural Sciences, The Biomedical Centre, SE-751 23 Uppsala, Sweden [3] Institute of Mathematics and Natural Sciences, Tallinn University, Narva mnt 25, EE-101 20 Tallinn, Estonia.
authors: Jan
authors: J
authors: Johansson
mesh_terms: D000818:Animals; D019281:Dimerization; D005349:Fibroins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D009682:Magnetic Resonance Spectroscopy; D047011:Silk / Q000096:biosynthesis* / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D013112:Spiders / Q000235:genetics / Q000378:metabolism*; D055672:Static Electricity
chemical_list: D047011:Silk; C086087:spidroin 1; D005349:Fibroins