Sammanfattning

Formation of spider silk from its constituent proteins-spidroins-involves changes from soluble helical/coil conformations to insoluble beta-sheet aggregates. This conversion needs to be regulated to avoid precocious aggregation proximally in the silk gland while still allowing rapid silk assembly in the distal parts. Lowering of pH from about 7 to 6 is apparently important for silk formation. The spidroin N-terminal domain (NT) undergoes stable dimerization and structural changes in this pH region, but the underlying mechanisms are incompletely understood. Here, we determine the NMR and crystal structures of Euprosthenops australis NT mutated in the dimer interface (A72R). Also, the NMR structure of wild-type (wt) E. australis NT at pH 7.2 and 300 mM sodium chloride was determined. The wt NT and A72R structures are monomers and virtually identical, but they differ from the subunit structure of dimeric wt NT mainly by having a tryptophan (W10) buried between helix 1 and helix 3, while W10 is surface exposed in the dimer. Wedging of the W10 side chain in monomeric NT tilts helix 3 approximately 5-6 angstrom into a position that is incompatible with that of the observed dimer structure. The structural differences between monomeric and dimeric NT domains explain the tryptophan fluorescence patterns of NT at pH 7 and pH 6 and indicate that the biological function of NT depends on conversion between the two conformations. (C) 2012 Elsevier Ltd. All rights reserved.

Nyckelord

NMR structure; X-ray structure; protein assembly; spidroin

Publicerad i

Journal of Molecular Biology
2012, Volym: 422, nummer: 4, sidor: 477-487
Utgivare: ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD

SLU författare

• Askarieh, Glareh

  • Institutionen för molekylärbiologi, Sveriges lantbruksuniversitet
    

• Hedhammar, My

  • Institutionen för husdjurens biovetenskaper, Sveriges lantbruksuniversitet
    

• Rising, Anna

  • Institutionen för husdjurens biovetenskaper, Sveriges lantbruksuniversitet
    

• Knight, Stefan David

  • Institutionen för molekylärbiologi, Sveriges lantbruksuniversitet
    

• Johansson, Jan

  • Institutionen för husdjurens biovetenskaper, Sveriges lantbruksuniversitet
    

UKÄ forskningsämne

Strukturbiologi


Publikationens identifierare

DOI: https://doi.org/10.1016/j.jmb.2012.06.004

Permanent länk till denna sida (URI)

https://res.slu.se/id/publ/42662