title: Novel Abetalipoproteinemia Missense Mutation Highlights the Importance of the N-Terminal β-Barrel in Microsomal Triglyceride Transfer Protein Function.
abstract: BACKGROUND
  The use of microsomal triglyceride transfer protein (MTP) inhibitors is limited to severe hyperlipidemias because of associated hepatosteatosis and gastrointestinal adverse effects. Comprehensive knowledge about the structure-function of MTP might help design new molecules that avoid steatosis. Characterization of mutations in MTP causing abetalipoproteinemia has revealed that the central α-helical and C-terminal β-sheet domains are important for protein disulfide isomerase binding and lipid transfer activity. Our aim was to identify and characterize mutations in the N-terminal domain to understand its function.
  
  
  RESULTS
  We identified a novel missense mutation (D169V) in a 4-month-old Turkish male child with severe signs of abetalipoproteinemia. To study the effect of this mutation on MTP function, we created mutants via site-directed mutagenesis. Although D169V was expressed in the endoplasmic reticulum and interacted with apolipoprotein B (apoB) 17, it was unable to bind protein disulfide isomerase, transfer lipids, and support apoB secretion. Computational modeling suggested that D169 could form an internal salt bridge with K187 and K189. Mutagenesis of these lysines to leucines abolished protein disulfide isomerase heterodimerization, lipid transfer, and apoB secretion, without affecting apoB17 binding. Furthermore, mutants with preserved charges (D169E, K187R, and K189R) rescued these activities.
  
  
  CONCLUSIONS
  D169V is detrimental because it disrupts an internal salt bridge leading to loss of protein disulfide isomerase binding and lipid transfer activities; however, it does not affect apoB binding. Thus, the N-terminal domain of MTP is also important for its lipid transfer activity.
authors.affiliation: From the School of Graduate Studies, Molecular and Cell Biology Program (M.T.W., J.J., J.S.), Department of Cell Biology (M.T.W., J.I., J.J., J.S., M.M.H.), Department of Pediatrics (M.M.H.), State University of New York Downstate Medical Center, Brooklyn, NY; Department of Life Sciences, University of Modena and Reggio Emilia, Modena, Italy (E.D.L., P.T.); Infancy Services, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (E.O); Department of Nutrition and Metabolism, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (M.G.); and Department of Research, VA New York Harbor Healthcare System, Brooklyn, NY (M.M.H.).
authors.forename: Meghan T
authors.initials: MT
authors.lastname: Walsh
authors.affiliation: From the School of Graduate Studies, Molecular and Cell Biology Program (M.T.W., J.J., J.S.), Department of Cell Biology (M.T.W., J.I., J.J., J.S., M.M.H.), Department of Pediatrics (M.M.H.), State University of New York Downstate Medical Center, Brooklyn, NY; Department of Life Sciences, University of Modena and Reggio Emilia, Modena, Italy (E.D.L., P.T.); Infancy Services, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (E.O); Department of Nutrition and Metabolism, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (M.G.); and Department of Research, VA New York Harbor Healthcare System, Brooklyn, NY (M.M.H.).
authors.forename: Jahangir
authors.initials: J
authors.lastname: Iqbal
authors.affiliation: From the School of Graduate Studies, Molecular and Cell Biology Program (M.T.W., J.J., J.S.), Department of Cell Biology (M.T.W., J.I., J.J., J.S., M.M.H.), Department of Pediatrics (M.M.H.), State University of New York Downstate Medical Center, Brooklyn, NY; Department of Life Sciences, University of Modena and Reggio Emilia, Modena, Italy (E.D.L., P.T.); Infancy Services, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (E.O); Department of Nutrition and Metabolism, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (M.G.); and Department of Research, VA New York Harbor Healthcare System, Brooklyn, NY (M.M.H.).
authors.forename: Joby
authors.initials: J
authors.lastname: Josekutty
authors.affiliation: From the School of Graduate Studies, Molecular and Cell Biology Program (M.T.W., J.J., J.S.), Department of Cell Biology (M.T.W., J.I., J.J., J.S., M.M.H.), Department of Pediatrics (M.M.H.), State University of New York Downstate Medical Center, Brooklyn, NY; Department of Life Sciences, University of Modena and Reggio Emilia, Modena, Italy (E.D.L., P.T.); Infancy Services, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (E.O); Department of Nutrition and Metabolism, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (M.G.); and Department of Research, VA New York Harbor Healthcare System, Brooklyn, NY (M.M.H.).
authors.forename: James
authors.initials: J
authors.lastname: Soh
authors.affiliation: From the School of Graduate Studies, Molecular and Cell Biology Program (M.T.W., J.J., J.S.), Department of Cell Biology (M.T.W., J.I., J.J., J.S., M.M.H.), Department of Pediatrics (M.M.H.), State University of New York Downstate Medical Center, Brooklyn, NY; Department of Life Sciences, University of Modena and Reggio Emilia, Modena, Italy (E.D.L., P.T.); Infancy Services, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (E.O); Department of Nutrition and Metabolism, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (M.G.); and Department of Research, VA New York Harbor Healthcare System, Brooklyn, NY (M.M.H.).
authors.forename: Enza
authors.initials: E
authors.lastname: Di Leo
authors.affiliation: From the School of Graduate Studies, Molecular and Cell Biology Program (M.T.W., J.J., J.S.), Department of Cell Biology (M.T.W., J.I., J.J., J.S., M.M.H.), Department of Pediatrics (M.M.H.), State University of New York Downstate Medical Center, Brooklyn, NY; Department of Life Sciences, University of Modena and Reggio Emilia, Modena, Italy (E.D.L., P.T.); Infancy Services, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (E.O); Department of Nutrition and Metabolism, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (M.G.); and Department of Research, VA New York Harbor Healthcare System, Brooklyn, NY (M.M.H.).
authors.forename: Eda
authors.initials: E
authors.lastname: Özaydin
authors.affiliation: From the School of Graduate Studies, Molecular and Cell Biology Program (M.T.W., J.J., J.S.), Department of Cell Biology (M.T.W., J.I., J.J., J.S., M.M.H.), Department of Pediatrics (M.M.H.), State University of New York Downstate Medical Center, Brooklyn, NY; Department of Life Sciences, University of Modena and Reggio Emilia, Modena, Italy (E.D.L., P.T.); Infancy Services, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (E.O); Department of Nutrition and Metabolism, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (M.G.); and Department of Research, VA New York Harbor Healthcare System, Brooklyn, NY (M.M.H.).
authors.forename: Mehmet
authors.initials: M
authors.lastname: Gündüz
authors.affiliation: From the School of Graduate Studies, Molecular and Cell Biology Program (M.T.W., J.J., J.S.), Department of Cell Biology (M.T.W., J.I., J.J., J.S., M.M.H.), Department of Pediatrics (M.M.H.), State University of New York Downstate Medical Center, Brooklyn, NY; Department of Life Sciences, University of Modena and Reggio Emilia, Modena, Italy (E.D.L., P.T.); Infancy Services, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (E.O); Department of Nutrition and Metabolism, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (M.G.); and Department of Research, VA New York Harbor Healthcare System, Brooklyn, NY (M.M.H.).
authors.forename: Patrizia
authors.initials: P
authors.lastname: Tarugi
authors.affiliation: From the School of Graduate Studies, Molecular and Cell Biology Program (M.T.W., J.J., J.S.), Department of Cell Biology (M.T.W., J.I., J.J., J.S., M.M.H.), Department of Pediatrics (M.M.H.), State University of New York Downstate Medical Center, Brooklyn, NY; Department of Life Sciences, University of Modena and Reggio Emilia, Modena, Italy (E.D.L., P.T.); Infancy Services, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (E.O); Department of Nutrition and Metabolism, Ankara Children's Health and Diseases Hematology-Oncology Training and Research Hospital, Ankara, Turkey (M.G.); and Department of Research, VA New York Harbor Healthcare System, Brooklyn, NY (M.M.H.). Mahmood.hussain@downstate.edu.
authors.forename: M Mahmood
authors.initials: MM
authors.lastname: Hussain
mesh_terms: D000012:Abetalipoproteinemia / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D001055:Apolipoproteins B / Q000378:metabolism; D001665:Binding Sites; D019556:COS Cells; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D002522:Chlorocebus aethiops; D004252:DNA Mutational Analysis; D004721:Endoplasmic Reticulum / Q000378:metabolism; D006801:Humans; D007223:Infant; D050356:Lipid Metabolism; D008297:Male; D020125:Mutation, Missense; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D014162:Transfection
chemical_list: D001055:Apolipoproteins B; D002352:Carrier Proteins; C106092:microsomal triglyceride transfer protein
keywords: apolipoproteins; apolipoproteins B; cardiovascular diseases; chylomicrons; lipids; lipoproteins; microsomal triglyceride transfer protein