title: Propofol shares the binding site with isoflurane and sevoflurane on leukocyte function-associated antigen-1.
abstract: BACKGROUND
  We previously demonstrated that propofol interacted with the leukocyte adhesion molecule leukocyte function-associated antigen-1 (LFA-1) and inhibited the production of interleukin-2 via LFA-1 in a dependent manner. However, the binding site(s) of propofol on LFA-1 remains unknown.
  
  
  METHODS
  First, the inhibition of LFA-1's ligand binding by propofol was confirmed in an enzyme-linked immunosorbent assay (ELISA) ELISA-type assay. The binding site of propofol on LFA-1 was probed with a photolabeling experiment using a photoactivatable propofol analog called azi-propofol-m. The adducted residues of LFA-1 by this compound were determined using liquid chromatography-mass spectrometry. In addition, the binding of propofol to the ligand-binding domain of LFA-1 was examined using 1-aminoanthracene (1-AMA) displacement assay. Furthermore, the binding site(s) of 1-AMA and propofol on LFA-1 was studied using the docking program GLIDE.
  
  
  RESULTS
  We demonstrated that propofol impaired the binding of LFA-1 to its ligand intercellular adhesion molecule-1. The photolabeling experiment demonstrated that the adducted residues were localized in the allosteric cavity of the ligand-binding domain of LFA-1 called "lovastatin site." The shift of fluorescence spectra was observed when 1-AMA was coincubated with the low-affinity conformer of LFA-1 ligand-binding domain (wild-type [WT] αL I domain), not with the high-affinity conformer, suggesting that 1-AMA bound only to WT αL I domain. In the 1-AMA displacement assay, propofol decreased 1-AMA fluorescence signal (at 520 nm), suggesting that propofol competed with 1-AMA and bound to the WT αL I domain. The docking simulation demonstrated that both 1-AMA and propofol bound to the lovastatin site, which agreed with the photolabeling experiment.
  
  
  CONCLUSIONS
  We demonstrated that propofol bound to the lovastatin site in LFA-1. Previously we showed that the volatile anesthetics isoflurane and sevoflurane bound to this site. Taken together, the lovastatin site is an example of the common binding sites for anesthetics currently used clinically.
authors: From the Department of Anesthesiology, Perioperative and Pain Medicine and Program in Cellular and Molecular Medicine, Boston Children's Hospital; Department of Anaesthesia, Harvard Medical School, Boston, Massachusetts; Department of Anesthesiology and Critical Care, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania; and Department of Molecular Pathobiology and Cell Adhesion Biology, Mie University, Graduate School of Medicine, Tsu, Mie, Japan.
authors: Koichi
authors: K
authors: Yuki
authors: Weiming
authors: W
authors: Bu
authors: Jin
authors: J
authors: Xi
authors: Motomu
authors: M
authors: Shimaoka
authors: Roderic
authors: R
authors: Eckenhoff
mesh_terms: D018685:Anesthetics, Inhalation / Q000378:metabolism*; D018686:Anesthetics, Intravenous / Q000378:metabolism*; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D018360:Crystallography, X-Ray; D006801:Humans; D007530:Isoflurane / Q000378:metabolism*; D008024:Ligands; D008148:Lovastatin / Q000378:metabolism; D016169:Lymphocyte Function-Associated Antigen-1 / Q000737:chemistry / Q000378:metabolism*; D008738:Methyl Ethers / Q000378:metabolism*; D015742:Propofol / Q000378:metabolism*; D011485:Protein Binding / Q000502:physiology; D000077149:Sevoflurane
chemical_list: D018685:Anesthetics, Inhalation; D018686:Anesthetics, Intravenous; D008024:Ligands; D016169:Lymphocyte Function-Associated Antigen-1; D008738:Methyl Ethers; D000077149:Sevoflurane; D008148:Lovastatin; D007530:Isoflurane; D015742:Propofol
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