title: 13C NMR of Nephila clavipes major ampullate silk gland.
abstract: The major ampullate glands of the spider Nephila clavipes contain approximately 0.2 microliter each of a highly concentrated (approximately 50%) solution of silk fibroin. Therefore, the reservoir of silk in these glands presents an ideal opportunity to observe prefolded conformations of a protein in its native state. To this end, the structure and conformation of major ampullate gland silk fibroin within the glands of the spider N. clavipes were examined by 13C NMR spectroscopy. These results were compared to those from silk protein first drawn from the spinneret and then denatured. The 13C NMR chemical shifts, along with infrared and circular dichroism data, suggest that the silk fibroin in the glands exists in dynamically averaged helical conformations. Furthermore, there is no evidence of proline residues in U-(13)C-D-glucose-labeled silk. This transient prefolded "molten fibril" state may correspond to the silk I form found in Bombyx mori silk. There is no evidence of the final beta-sheet structure in the ampullate gland silk fibroin before final silk processing. However, the conformation of silk in the glands appears to be in a highly metastable state, as plasticization with water produces the beta-sheet structure. Therefore, the ducts connecting the ampullate glands to the spinnerets play a larger role in silk processing than previously thought.
authors.affiliation: Center for Advanced Technology in Biotechnology, Cornell University, Ithaca, New York 14853, USA.
authors.forename: D H
authors.initials: DH
authors.lastname: Hijirida
authors.forename: K G
authors.initials: KG
authors.lastname: Do
authors.forename: C
authors.initials: C
authors.lastname: Michal
authors.forename: S
authors.initials: S
authors.lastname: Wong
authors.forename: D
authors.initials: D
authors.lastname: Zax
authors.forename: L W
authors.initials: LW
authors.lastname: Jelinski
mesh_terms: D000409:Alanine; D000818:Animals; D002247:Carbon Isotopes; D002942:Circular Dichroism; D005612:Freeze Drying; D019476:Insect Proteins / Q000032:analysis* / Q000096:biosynthesis; D009682:Magnetic Resonance Spectroscopy; D012627:Sebaceous Glands / Q000648:ultrastructure*; D047011:Silk; D017550:Spectroscopy, Fourier Transform Infrared; D013112:Spiders
chemical_list: D002247:Carbon Isotopes; D019476:Insect Proteins; D047011:Silk; D000409:Alanine