A model for the structure of the C-terminal domain of dragline spider silk and the role of its conserved cysteine.
Dragline spider silk fibers have extraordinary attributes as biomaterials of superior strength and toughness. Previously we have shown that the conserved C-terminal domain of a dragline spider silk protein is necessary for directing oriented microfiber formation. Here we present for the first time a state-of-the-art model of the three-dimensional structure of this domain, and, by comparing several dragline proteins, identify its key evolutionarily conserved features. Further, using the baculovirus expression system, we produced recombinant proteins that are mutated in the unique cysteine residue present in the domain. While a conservative mutation to serine allows fiber formation, thus demonstrating that there is no need for disulfide bond formation in this system, a mutation to arginine significantly alters the local surface properties, preventing fiber formation. These experimental results are in agreement with our model, wherein the cysteine is localized in a highly conserved hydrophobic loop that we predict to be important for the protein-protein interactions of this domain and hence also for fiber formation.
Ittah
Shmulik
S
Department of Cell and Animal Biology, Silberman Life Sciences Institute, Edmond Safra Campus at Givat-Ram, The Hebrew University, Jerusalem 91904, Israel.
Michaeli
Amit
A
Goldblum
Amiram
A
Gat
Uri
U
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D017124:Conserved Sequence; D003545:Cysteine / Q000737:chemistry* / Q000378:metabolism*; D005786:Gene Expression Regulation; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009036:Moths; D017434:Protein Structure, Tertiary; D047011:Silk / Q000737:chemistry*; D013112:Spiders / Q000737:chemistry*
D047011:Silk; D003545:Cysteine