Barnes S

References (6)

Title : The Cystic Fibrosis Transmembrane Conductance Regulator Potentiator Ivacaftor Augments Mucociliary Clearance Abrogating Cystic Fibrosis Transmembrane Conductance Regulator Inhibition by Cigarette Smoke - Raju_2017_Am.J.Respir.Cell.Mol.Biol_56_99
Author(s) : Raju SV , Lin VY , Liu L , McNicholas CM , Karki S , Sloane PA , Tang L , Jackson PL , Wang W , Wilson L , Macon KJ , Mazur M , Kappes JC , DeLucas LJ , Barnes S , Kirk K , Tearney GJ , Rowe SM
Ref : American Journal of Respiratory Cellular & Molecular Biology , 56 :99 , 2017
Abstract : Acquired cystic fibrosis transmembrane conductance regulator (CFTR) dysfunction may contribute to chronic obstructive pulmonary disease pathogenesis and is a potential therapeutic target. We sought to determine the acute effects of cigarette smoke on ion transport and the mucociliary transport apparatus, their mechanistic basis, and whether deleterious effects could be reversed with the CFTR potentiator ivacaftor (VX-770). Primary human bronchial epithelial (HBE) cells and human bronchi were exposed to cigarette smoke extract (CSE) and/or ivacaftor. CFTR function and expression were measured in Ussing chambers and by surface biotinylation. CSE-derived acrolein modifications on CFTR were determined by mass spectroscopic analysis of purified protein, and the functional microanatomy of the airway epithelia was measured by 1-mum resolution optical coherence tomography. CSE reduced CFTR-dependent current in HBE cells (P < 0.05) and human bronchi (P < 0.05) within minutes of exposure. The mechanism involved CSE-induced reduction of CFTR gating, decreasing CFTR open-channel probability by approximately 75% immediately after exposure (P < 0.05), whereas surface CFTR expression was partially reduced with chronic exposure, but was stable acutely. CSE treatment of purified CFTR resulted in acrolein modifications on lysine and cysteine residues that likely disrupt CFTR gating. In primary HBE cells, CSE reduced airway surface liquid depth (P < 0.05) and ciliary beat frequency (P < 0.05) within 60 minutes that was restored by coadministration with ivacaftor (P < 0.005). Cigarette smoking transmits acute reductions in CFTR activity, adversely affecting the airway surface. These effects are reversible by a CFTR potentiator in vitro, representing a potential therapeutic strategy in patients with chronic obstructive pulmonary disease with chronic bronchitis.
ESTHER : Raju_2017_Am.J.Respir.Cell.Mol.Biol_56_99
PubMedSearch : Raju_2017_Am.J.Respir.Cell.Mol.Biol_56_99
PubMedID: 27585394

Title : Rat liver bile acid CoA:amino acid N-acyltransferase: expression, characterization, and peroxisomal localization - He_2003_J.Lipid.Res_44_2242
Author(s) : He D , Barnes S , Falany CN
Ref : J Lipid Res , 44 :2242 , 2003
Abstract : Bile acid CoA:amino acid N-acyltransferase (BAT) is responsible for the amidation of bile acids with the amino acids taurine and glycine. Rat liver BAT (rBAT) cDNA was isolated from a rat liver lambdaZAP cDNA library and expressed in Sf9 insect cells using a baculoviral vector. rBAT displayed 65% amino acid sequence homology with human BAT (hBAT) and 85% homology with mouse BAT (mBAT). Similar to hBAT, expressed rBAT was capable of forming both taurine and glycine conjugates with cholyl-CoA. mBAT, which is highly homologous to rBAT, forms only taurine conjugated bile acids (Falany, C. N., H. Fortinberry, E. H. Leiter, and S. Barnes. 1997. Cloning and expression of mouse liver bile acid CoA: Amino acid N-acyltransferase. J. Lipid Res. 38: 86-95). Immunoblot analysis of rat tissues detected rBAT only in rat liver cytosol following homogenization and ultracentrifugation. Subcellular localization of rBAT detected activity and immunoreactive protein in both cytosol and isolated peroxisomes. Rat bile acid CoA ligase (rBAL), the enzyme responsible for the formation of bile acid CoA esters, was detected only in rat liver microsomes. Treatment of rats with clofibrate, a known peroxisomal proliferator, significantly induced rBAT activity, message, and immunoreactive protein in rat liver. Peroxisomal membrane protein-70, a marker for peroxisomes, was also induced by clofibrate, whereas rBAL activity and protein amount were not affected. In summary, rBAT is capable of forming both taurine and glycine bile acid conjugates and the enzyme is localized primarily in peroxisomes in rat liver.
ESTHER : He_2003_J.Lipid.Res_44_2242
PubMedSearch : He_2003_J.Lipid.Res_44_2242
PubMedID: 12951368
Gene_locus related to this paper: ratno-BAAT

Title : Conserved residues in the putative catalytic triad of human bile acid Coenzyme A:amino acid N-acyltransferase - Sfakianos_2002_J.Biol.Chem_277_47270
Author(s) : Sfakianos MK , Wilson L , Sakalian M , Falany CN , Barnes S
Ref : Journal of Biological Chemistry , 277 :47270 , 2002
Abstract : Human bile acid-CoA:amino acid N-acyltransferase (hBAT), an enzyme catalyzing the conjugation of bile acids with the amino acids glycine or taurine has significant sequence homology with dienelactone hydrolases and other alpha/beta hydrolases. These enzymes have a conserved catalytic triad that maps onto the mammalian BATs at residues Cys-235, Asp-328, and His-362 of the human sequence, albeit that the hydrolases contain a serine instead of a cysteine. In the present study, the function of the putative catalytic triad of hBAT was examined by chemical modification with the cysteine alkylating reagent N-ethylmaleimide (NEM) and by site-directed mutagenesis of the triad residues followed by enzymology studies of mutant and wild-type hBATs. Treatment with NEM caused inactivation of wild-type hBAT. However, preincubation of wild-type hBAT with the substrate cholyl-CoA before NEM treatment prevented loss of N-acyltransferase activity. Substitution of His-362 or Asp-328 with alanine results in inactivation of hBAT. Although substitution of Cys-235 with serine generated an hBAT mutant with lower N-acyltransferase activity, it substantially increased the bile acid-CoA thioesterase activity compared with wild type. In summary, data from this study support the existence of an essential catalytic triad within hBAT consisting of Cys-235, His-362, and Asp-328 with Cys-235 serving as the probable nucleophile and thus the site of covalent attachment of the bile acid molecule.
ESTHER : Sfakianos_2002_J.Biol.Chem_277_47270
PubMedSearch : Sfakianos_2002_J.Biol.Chem_277_47270
PubMedID: 12239217
Gene_locus related to this paper: human-BAAT

Title : Cloning, expression, and chromosomal localization of mouse liver bile acid CoA:amino acid N-acyltransferase - Falany_1997_J.Lipid.Res_38_1139
Author(s) : Falany CN , Fortinberry H , Leiter EH , Barnes S
Ref : J Lipid Res , 38 :1139 , 1997
Abstract : A mouse liver lambda Zap XR cDNA library was screened using the coding region of human bile acid CoA:amino acid N-acyltransferase (BAT) cDNA as a probe. Ten positive clones were isolated and purified, two of which apparently possessed complete open reading frames for BAT based on sequence analysis of the ends of the cDNAs. One clone (mBAT#9) was selected for sequence analysis and characterization. mBAT#9 is 1869 basepairs in length and the full-length cDNA possesses a 189 basepair 5'-nontranslated region, an open-reading frame of 1260 basepairs, and a 404 basepair 3'-nontranslated region followed by a poly(A) tail. The open-reading frame codes for a 420 amino acid protein with a calculated molecular mass of 46,525 daltons. The structural gene for mBAT was mapped to mouse Chromosome 4. The amino acid sequence of mBAT is 69% identical and 84% similar to that of hBAT, and 86% identical and 95% similar to that of kan-1, a putative rat liver BAT. Enzymatically active mBAT was expressed in E. coli using the bacterial expression vector pKK233-2. Immunoblot analysis of expressed mBAT with rabbit anti-human BAT polyclonal antibodies detected a single protein with a molecular mass of approximately 45,000 daltons. Cytosol from cells transformed with mBAT#9/pKK233-2 possessed significant amounts of BAT-catalyzed conjugating activity with taurine as substrate but the expressed enzyme did not use glycine or fluoro-beta-alanine as substrates. The K(m) value for taurine was 1.9 mM +/- 0.1 mM in reactions with cholyl CoA as a cosubstrate. The specificity of mBAT for taurine as a substrate was confirmed by the demonstration, using HPLC-electrospray ionization mass spectrometry, that mouse gallbladder bile contained only taurine conjugates of bile acids. The identification of the types of amino acid conjugates of bile acids present in mouse bile had not been previously reported. These results indicate that a taurine-specific form of BAT has been cloned and expressed from mouse liver.
ESTHER : Falany_1997_J.Lipid.Res_38_1139
PubMedSearch : Falany_1997_J.Lipid.Res_38_1139
PubMedID: 9215542
Gene_locus related to this paper: mouse-BAAT

Title : Glycine and taurine conjugation of bile acids by a single enzyme. Molecular cloning and expression of human liver bile acid CoA:amino acid N-acyltransferase. -
Author(s) : Falany CN , Johnson MR , Barnes S , Diasio RB
Ref : Journal of Biological Chemistry , 269 :19375 , 1994
PubMedID: 8034703
Gene_locus related to this paper: human-BAAT

Title : Purification and characterization of bile acid-CoA:amino acid N-acyltransferase from human liver - Johnson_1991_J.Biol.Chem_266_10227
Author(s) : Johnson MR , Barnes S , Kwakye JB , Diasio RB
Ref : Journal of Biological Chemistry , 266 :10227 , 1991
Abstract : The bile acid-conjugating enzyme, bile acid-CoA: amino acid N-acyltransferase, was purified 480-fold from the soluble fraction of homogenized frozen human liver. Purification was accomplished by a combination of anion exchange chromatography, chromatofocusing, glycocholate-AH-Sepharose affinity chromatography, and high performance liquid chromatography (HPLC) gel filtration. Following purification, the reduced, denatured enzyme migrated as a single 50-kDa protein band by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A similar molecular mass was obtained for the native enzyme by HPLC gel filtration. Elution from the chromatofocusing column suggested an apparent isoelectric point of 6.0 (+/- 0.2). Using a rabbit polyclonal antibody raised against the purified enzyme, Western blot analysis using 100,000 x g human liver supernatant confirmed that the affinity-purified polyclonal antibody was specific for human liver bile acid-CoA:amino acid N-acyltransferase. The purified enzyme utilized glycine, taurine, and 2-fluoro-beta-alanine (a 5-fluorouracil catabolite), but not beta-alanine, as substrates. Kinetic studies revealed apparent Km values for taurine, 2-fluoro-beta-alanine, and glycine of 1.1, 2.2, and 5.8 mM, respectively, with corresponding Vmax values of 0.33, 0.19, and 0.77 mumol/min/mg protein. These data demonstrate that a single monomeric enzyme is responsible for the conjugation of bile acids with glycine or taurine in human liver.
ESTHER : Johnson_1991_J.Biol.Chem_266_10227
PubMedSearch : Johnson_1991_J.Biol.Chem_266_10227
PubMedID: 2037576
Gene_locus related to this paper: human-BAAT