title: Solution structure of the heterotrimeric complex between the interaction domains of RFX5 and RFXAP from the RFX gene regulatory complex.
abstract: The mammalian immune response is mediated by a heterotetrameric transcriptional control complex, called regulatory factor X (RFX), that regulates the expression of major histocompatibility complex class II genes. RFX comprises three proteins: RFX5 (two copies), RFXAP, and RFXB, and mutations and deletions that prevent the assembly of the RFX complex have been linked to a severe immunodeficiency disorder. Two RFX5 molecules and one RFXAP molecule assemble in the cytoplasm prior to nuclear localization, a process mediated by an N-terminal "dimerization domain" of RFX5 (RFX5(N)) and a C-terminal domain of RFXAP (RFXAP(C)). We previously presented evidence that RFXAP(C) is unstructured in the absence of RFX5(N) but adopts a regular structure in the RFX5(N)(2)-RFXAP(C) complex and that the RFX5(N)(2)-RFXAP(C) complex binds RFXB with high affinity. We now report the structure of the RFX5(N)(2)-RFXAP(C) complex, determined in solution by (15)N- and (13)C-edited NMR spectroscopy. RFX5(N) consists of a long central helix flanked by two shorter helices. The central helices of the two RFX5(N) molecules form an antiparallel coiled coil, and the flanking helices pack at the ends of the long helices in a perpendicular arrangement such that the RFX5(N) dimer is shaped like a staple. RFXAP(C) consists of two α-helices that form a V-shaped structure that packs within the RFX5(N)(2) staple. Leucine residues in the leucine-rich region of RFX5(N) (62-LYLYLQL-68) that are critical for major histocompatibility complex class II gene expression in vivo contribute to both the dimer (Leu64 and Leu68) and the RFX5(N)-RFXAP(C) interfaces (Leu62 and Leu66). The clustering of hydrophobic residues from different regions of RFXAP(C) suggests a potential binding site for RFXB.
authors: Department of Chemistry and Biochemistry, University of Maryland, Baltimore County, 1000 Hilltop Circle, Baltimore, MD 21250, USA.
authors: Kholiswa M
authors: KM
authors: Laird
authors: Department of Chemistry and Biochemistry, University of Maryland, Baltimore County, 1000 Hilltop Circle, Baltimore, MD 21250, USA.
authors: LaTese L
authors: LL
authors: Briggs
authors: Department of Microbiology and Immunology, Emory University School of Medicine, 1510 Clifton Road, Atlanta, GA 30322, USA.
authors: Jeremy M
authors: JM
authors: Boss
authors: Department of Chemistry and Biochemistry, University of Maryland, Baltimore County, 1000 Hilltop Circle, Baltimore, MD 21250, USA.
authors: Michael F
authors: MF
authors: Summers
authors: Department of Chemistry and Biochemistry, University of Maryland, Baltimore County, 1000 Hilltop Circle, Baltimore, MD 21250, USA. Electronic address: colin.garvie@roche.com.
authors: Colin W
authors: CW
authors: Garvie
mesh_terms: D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D000071701:Regulatory Factor X Transcription Factors; D014157:Transcription Factors / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D004268:DNA-Binding Proteins; C093524:RFX5 protein, human; C413004:RFXAP protein, human; D000071701:Regulatory Factor X Transcription Factors; D014157:Transcription Factors