title: Recombinant human PPAR-beta/delta ligand-binding domain is locked in an activated conformation by endogenous fatty acids.
abstract: High-resolution crystallographic structures of recombinant human peroxisome proliferator-activated receptor ligand-binding domain (isotype beta/delta) reveal a fatty acid in the binding site. Mass spectrometry confirmed the presence of C16:0, C16:1, C18:0 and C18:1 in a ratio of approximately 3:2:1:4 with 11, Z-octadecenoic acid (cis-vaccenic acid) identified as the predominant species. These are endogenous fatty acids acquired from the bacterial expression system, and serve to lock the ligand-binding domain into the activated conformation. A requirement for crystal growth, the additive n-heptyl-beta-d-glucopyranoside, binds near the activation function helix where recognition of co-activator proteins occurs. Our observations suggest potential physiological ligands for human PPAR-beta/delta and highlight that reported binding studies must be treated with caution unless endogenous fatty acids have been removed from the sample prior to analysis.
authors.affiliation: Division of Biological Chemistry and Molecular Microbiology, School of Life Sciences, University of Dundee, Dundee DD1 5EH, UK.
authors.forename: Stewart A
authors.initials: SA
authors.lastname: Fyffe
authors.forename: Magnus S
authors.initials: MS
authors.lastname: Alphey
authors.forename: Lori
authors.initials: L
authors.lastname: Buetow
authors.forename: Terry K
authors.initials: TK
authors.lastname: Smith
authors.forename: Michael A J
authors.initials: MA
authors.lastname: Ferguson
authors.forename: Morten D
authors.initials: MD
authors.lastname: Sørensen
authors.forename: Fredrik
authors.initials: F
authors.lastname: Björkling
authors.forename: William N
authors.initials: WN
authors.lastname: Hunter
mesh_terms: D018360:Crystallography, X-Ray; D004789:Enzyme Activation; D005227:Fatty Acids / Q000737:chemistry* / Q000378:metabolism; D005936:Glucans / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D007527:Isoenzymes / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D047494:PPAR delta / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D048068:PPAR-beta / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D005227:Fatty Acids; D005936:Glucans; D007527:Isoenzymes; D047494:PPAR delta; D048068:PPAR-beta; D011994:Recombinant Proteins