title: High throughput protein fold identification by using experimental constraints derived from intramolecular cross-links and mass spectrometry.
abstract: We have used intramolecular cross-linking, MS, and sequence threading to rapidly identify the fold of a model protein, bovine basic fibroblast growth factor (FGF)-2. Its tertiary structure was probed with a lysine-specific cross-linking agent, bis(sulfosuccinimidyl) suberate (BS(3)). Sites of cross-linking were determined by tryptic peptide mapping by using time-of-flight MS. Eighteen unique intramolecular lysine (Lys-Lys) cross-links were identified. The assignments for eight cross-linked peptides were confirmed by using post source decay MS. The interatomic distance constraints were all consistent with the tertiary structure of FGF-2. These relatively few constraints, in conjunction with threading, correctly identified FGF-2 as a member of the beta-trefoil fold family. To further demonstrate utility, we used the top-scoring homolog, IL-1beta, to build an FGF-2 homology model with a backbone error of 4.8 A (rms deviation). This method is fast, is general, uses small amounts of material, and is amenable to automation.
authors.affiliation: Department of Pharmaceutical Chemistry, University of California, San Francisco, CA 94143-0446, USA.
authors.forename: M M
authors.initials: MM
authors.lastname: Young
authors.forename: N
authors.initials: N
authors.lastname: Tang
authors.forename: J C
authors.initials: JC
authors.lastname: Hempel
authors.forename: C M
authors.initials: CM
authors.lastname: Oshiro
authors.forename: E W
authors.initials: EW
authors.lastname: Taylor
authors.forename: I D
authors.initials: ID
authors.lastname: Kuntz
authors.forename: B W
authors.initials: BW
authors.lastname: Gibson
authors.forename: G
authors.initials: G
authors.lastname: Dollinger
mesh_terms: D000818:Animals; D002417:Cattle; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D003432:Cross-Linking Reagents / Q000494:pharmacology*; D016222:Fibroblast Growth Factor 2 / Q000737:chemistry* / Q000145:classification / Q000187:drug effects; D007375:Interleukin-1 / Q000737:chemistry; D008958:Models, Molecular; D017510:Protein Folding; D017434:Protein Structure, Tertiary / Q000187:drug effects; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013388:Succinimides / Q000494:pharmacology*
chemical_list: D003432:Cross-Linking Reagents; D007375:Interleukin-1; D013388:Succinimides; D016222:Fibroblast Growth Factor 2; C035760:bis(sulfosuccinimidyl)suberate