title: Biophysical and structural characterization of the thioredoxin-binding domain of protein kinase ASK1 and its interaction with reduced thioredoxin.
abstract: Apoptosis signal-regulating kinase 1 (ASK1), a mitogen-activated protein kinase kinase kinase, plays a key role in the pathogenesis of multiple diseases. Its activity is regulated by thioredoxin (TRX1) but the precise mechanism of this regulation is unclear due to the lack of structural data. Here, we performed biophysical and structural characterization of the TRX1-binding domain of ASK1 (ASK1-TBD) and its complex with reduced TRX1. ASK1-TBD is a monomeric and rigid domain that forms a stable complex with reduced TRX1 with 1:1 molar stoichiometry. The binding interaction does not involve the formation of intermolecular disulfide bonds. Residues from the catalytic WCGPC motif of TRX1 are essential for complex stability with Trp(31) being directly involved in the binding interaction as suggested by time-resolved fluorescence. Small-angle x-ray scattering data reveal a compact and slightly asymmetric shape of ASK1-TBD and suggest reduced TRX1 interacts with this domain through the large binding interface without inducing any dramatic conformational change.
authors.affiliation: From the Department of Physical and Macromolecular Chemistry, Faculty of Science, Charles University in Prague, 12843 Prague, the Institute of Physiology, Academy of Sciences of the Czech Republic, 14220 Prague, and.
authors.forename: Dalibor
authors.initials: D
authors.lastname: Kosek
authors.affiliation: From the Department of Physical and Macromolecular Chemistry, Faculty of Science, Charles University in Prague, 12843 Prague, the Institute of Physiology, Academy of Sciences of the Czech Republic, 14220 Prague, and.
authors.forename: Salome
authors.initials: S
authors.lastname: Kylarova
authors.affiliation: From the Department of Physical and Macromolecular Chemistry, Faculty of Science, Charles University in Prague, 12843 Prague, the Institute of Physiology, Academy of Sciences of the Czech Republic, 14220 Prague, and.
authors.forename: Katarina
authors.initials: K
authors.lastname: Psenakova
authors.affiliation: From the Department of Physical and Macromolecular Chemistry, Faculty of Science, Charles University in Prague, 12843 Prague.
authors.forename: Lenka
authors.initials: L
authors.lastname: Rezabkova
authors.affiliation: the Institute of Physics, Faculty of Mathematics and Physics, Charles University in Prague, 12116 Prague, Czech Republic.
authors.forename: Petr
authors.initials: P
authors.lastname: Herman
authors.affiliation: the Institute of Physics, Faculty of Mathematics and Physics, Charles University in Prague, 12116 Prague, Czech Republic.
authors.forename: Jaroslav
authors.initials: J
authors.lastname: Vecer
authors.affiliation: the Institute of Physiology, Academy of Sciences of the Czech Republic, 14220 Prague, and.
authors.forename: Veronika
authors.initials: V
authors.lastname: Obsilova
authors.affiliation: From the Department of Physical and Macromolecular Chemistry, Faculty of Science, Charles University in Prague, 12843 Prague, the Institute of Physiology, Academy of Sciences of the Czech Republic, 14220 Prague, and obsil@natur.cuni.cz.
authors.forename: Tomas
authors.initials: T
authors.lastname: Obsil
mesh_terms: D001703:Biophysics; D002942:Circular Dichroism; D048848:MAP Kinase Kinase Kinase 5 / Q000378:metabolism*; D010084:Oxidation-Reduction; D011485:Protein Binding; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence; D013879:Thioredoxins / Q000378:metabolism*; D014461:Ultracentrifugation
chemical_list: D013879:Thioredoxins; D048848:MAP Kinase Kinase Kinase 5
keywords: Analytical Ultracentrifugation; Apoptosis Signal-regulating Kinase 1 (ASK1); Circular Dichroism (CD); Fluorescence; Small-angle X-ray Scattering (SAXS); Thioredoxin