title: Elucidating metabolic pathways for amino acid incorporation into dragline spider silk using 13C enrichment and solid state NMR.
abstract: Spider silk has been evolutionarily optimized for contextual mechanical performance over the last 400 Ma. Despite precisely balanced mechanical properties, which have yet to be reproduced, the underlying molecular architecture of major ampullate spider silk can be simplified being viewed as a versatile block copolymer. Four primary amino acid motifs: polyalanine, (GA)(n), GPGXX, and GGX (X = G,A,S,Q,L,Y) will be considered in this study. Although synthetic mimetics of many of these amino acid motifs have been produced in several biological systems, the source of spider silk's mechanical integrity remains elusive. Mechanical robustness may be a product not only of the amino acid structure but also of the tertiary structure of the silk. Historically, solid state nuclear magnetic resonance (ssNMR) has been used to reveal the crystalline structure of the polyalanine motif; however, limitations in amino acid labeling techniques have obscured the structures of the GGX and GPGXX motifs thought to be responsible for the structural mobility of spider silk. We describe the use of metabolic pathways to label tyrosine for the first time as well as to improve the labeling efficiency of proline. These improved labeling techniques will allow the previously unknown tertiary structures of major ampullate silk to be probed.
authors.affiliation: Department of Molecular Biology, University of Wyoming, 16th and Gibbon, Laramie, WY 82070, USA. melinda.creager.phd@gmail.com
authors.forename: Melinda S
authors.initials: MS
authors.lastname: Creager
authors.forename: Thomas
authors.initials: T
authors.lastname: Izdebski
authors.forename: Amanda E
authors.initials: AE
authors.lastname: Brooks
authors.forename: Randolph V
authors.initials: RV
authors.lastname: Lewis
mesh_terms: D000596:Amino Acids / Q000378:metabolism*; D000818:Animals; D002247:Carbon Isotopes; D005260:Female; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D047011:Silk / Q000737:chemistry / Q000378:metabolism*; D013045:Species Specificity; D013112:Spiders
chemical_list: D000596:Amino Acids; D002247:Carbon Isotopes; D047011:Silk