title: Characterization and binding specificity of the monomeric STAT3-SH2 domain.
abstract: Signal transducers and activators of transcription (STATs) are important mediators of cytokine signal transduction. STAT factors are recruited to phosphotyrosine-containing motifs of activated receptor chains via their SH2 domains. The subsequent tyrosine phosphorylation of the STATs leads to their dissociation from the receptor, dimerization, and translocation to the nucleus. Here we describe the expression, purification, and refolding of the STAT3-SH2 domain. Proper folding of the isolated protein was proven by circular dichroism and fluorescence spectroscopy. The STAT3-SH2 domain undergoes a conformational change upon dimerization. Using an enzyme-linked immunosorbent assay we demonstrate that the monomeric domain binds to specific phosphotyrosine peptides. The specificity of binding to phosphotyrosine peptides was assayed with the tyrosine motif encompassing Tyr705 of STAT3 and with all tyrosine motifs present in the cytoplasmic tail of the signal transducer gp130.
authors.affiliation: Institut für Biochemie, Rheinisch-Westfälische Technische Hochschule Aachen, Pauwelsstrasse 30, D-52074 Aachen, Germany.
authors.forename: S
authors.initials: S
authors.lastname: Haan
authors.forename: U
authors.initials: U
authors.lastname: Hemmann
authors.forename: U
authors.initials: U
authors.lastname: Hassiepen
authors.forename: F
authors.initials: F
authors.lastname: Schaper
authors.forename: J
authors.initials: J
authors.lastname: Schneider-Mergener
authors.forename: A
authors.initials: A
authors.lastname: Wollmer
authors.forename: P C
authors.initials: PC
authors.lastname: Heinrich
authors.forename: J
authors.initials: J
authors.lastname: Grötzinger
mesh_terms: D001665:Binding Sites; D002942:Circular Dichroism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli; D019000:Phosphotyrosine / Q000378:metabolism; D011487:Protein Conformation; D017510:Protein Folding; D050796:STAT3 Transcription Factor; D013050:Spectrometry, Fluorescence; D015534:Trans-Activators / Q000378:metabolism*; D018909:src Homology Domains
chemical_list: D004268:DNA-Binding Proteins; D050796:STAT3 Transcription Factor; D015534:Trans-Activators; D019000:Phosphotyrosine