title: Automated resonance assignment of the 21kDa stereo-array isotope labeled thioldisulfide oxidoreductase DsbA.
abstract: The automated chemical shift assignment algorithm FLYA has been extended for use with stereo-array isotope labeled (SAIL) proteins to determine the sequence-specific resonance assignments of large proteins. Here we present the assignment of the backbone and sidechain chemical shifts of the 21kDa thioldisulfide oxidoreductase DsbA from Escherichia coli that were determined with the SAIL-FLYA algorithm in conjunction with automated peak picking. No manual corrections of peak lists or assignments were applied. The assignments agreed with manually determined reference assignments in 95.4% of the cases if 16 input spectra were used, 94.1% if only 3D13C/15N-resolved NOESY, CBCA(CO)NH, and 2D [13C/15N,1H]-HSQC were used, and 86.8% if exclusively 3D13C/15N-resolved NOESY spectra were used. Considering only the assignments that are classified as reliable by the SAIL-FLYA algorithm, the degrees of agreement increased to 97.5%, 96.5%, and 94.2%, respectively. With our approach it is thus possible to automatically obtain almost complete and correct assignments of proteins larger than 20kDa.
authors.affiliation: Institute of Biophysical Chemistry, Center for Biomolecular Magnetic Resonance, and Frankfurt Institute for Advanced Studies, Goethe University Frankfurt am Main, Max-von-Laue-Str. 9, 60438 Frankfurt am Main, Germany.
authors.forename: Elena
authors.initials: E
authors.lastname: Schmidt
authors.affiliation: Department of Chemistry, Graduate School of Science and Engineering, Tokyo Metropolitan University, Hachioji, Tokyo 192-0397, Japan.
authors.forename: Teppei
authors.initials: T
authors.lastname: Ikeya
authors.affiliation: Structural Biology Research Center, Graduate School of Science, Nagoya University, Nagoya 464-8602, Japan.
authors.forename: Mitsuhiro
authors.initials: M
authors.lastname: Takeda
authors.affiliation: Institute of Biophysical Chemistry, Center for Biomolecular Magnetic Resonance, and Frankfurt Institute for Advanced Studies, Goethe University Frankfurt am Main, Max-von-Laue-Str. 9, 60438 Frankfurt am Main, Germany.
authors.forename: Frank
authors.initials: F
authors.lastname: Löhr
authors.affiliation: Institute of Biophysical Chemistry, Center for Biomolecular Magnetic Resonance, and Frankfurt Institute for Advanced Studies, Goethe University Frankfurt am Main, Max-von-Laue-Str. 9, 60438 Frankfurt am Main, Germany.
authors.forename: Lena
authors.initials: L
authors.lastname: Buchner
authors.affiliation: Department of Chemistry, Graduate School of Science and Engineering, Tokyo Metropolitan University, Hachioji, Tokyo 192-0397, Japan.
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Ito
authors.affiliation: Department of Chemistry, Graduate School of Science and Engineering, Tokyo Metropolitan University, Hachioji, Tokyo 192-0397, Japan; Structural Biology Research Center, Graduate School of Science, Nagoya University, Nagoya 464-8602, Japan. Electronic address: kainosho@tmu.ac.jp.
authors.forename: Masatsune
authors.initials: M
authors.lastname: Kainosho
authors.affiliation: Institute of Biophysical Chemistry, Center for Biomolecular Magnetic Resonance, and Frankfurt Institute for Advanced Studies, Goethe University Frankfurt am Main, Max-von-Laue-Str. 9, 60438 Frankfurt am Main, Germany; Department of Chemistry, Graduate School of Science and Engineering, Tokyo Metropolitan University, Hachioji, Tokyo 192-0397, Japan. Electronic address: guentert@em.uni-frankfurt.de.
authors.forename: Peter
authors.initials: P
authors.lastname: Güntert
keywords: Automated assignment; CYANA; Disulfide bond; FLYA; SAIL; Thioldisulfide oxidoreductase