Affinity Chromatography of Neuropathy Target Esterase

Details

• Personal Author:
  Hammock BD ; McNamee MG ; Szekacs A ; Thomas TC ; Wilson BW
• Description:
  Neuropathy target esterase (NTE) is a membrane-bound protein which has been proposed as the target site in nerve tissue for initiation of organophosphate induced delayed neuropathy (OPIDN). Efforts to characterize NTE and to determine the mechanism of its involvement in OPIDN have been hampered by the lack of a suitable method for its purification. We describe here the development of a trifluoromethyl ketone liganded affinity gel which selectively binds NTE. Triton X-100/NaCl extracts of NTE from chick embryo brain microsomal membranes were adsorbed to an affinity gel prepared by attachment of 3(9'-mercaptononylthio)-1,1,1-trifluoropropan-2-one to epoxy-activated Sepharose CL4B (MNTFP-Sepharose). Typically 70-80% of NTE activity is bound under conditions in which undetectable quantities of total protein bound (< 4%). It proved difficult to elute active NTE under non-denaturing conditions, but SDS-PAGE analysis of MNTFP-Sepharose bound proteins eluted with 2% SDS identified a 155 kDa NTE-like protein that bound in a trifluoromethylketone- or mipafox-sensitive but paraoxon-insensitive manner. The levels of inhibition of binding correlated with the inhibition of activity and suggested that the 155-kDa band was composed of a single protein. MNTFP-Sepharose affinity chromatography in combination with preparative SDS-PAGE therefore holds promise as a method for obtaining microgram quantities of NTE for chemical analysis and sequencing. [Description provided by NIOSH]
• Subjects:
  Butanones Chemistry Chromatography Energy Metabolism Filtration Nervous System Occupational Health Organophosphates Safety
• Keywords:
  Chemical-analysis Chromatographic-analysis Ketones Metabolic-study Molecular-structure Nerve-tissue Nerves Neuropathy Organo-phosphorus-compounds
• ISSN:
  0009-2797
• Document Type:
  Text
• Funding:
  Cooperative Agreement
• Genre:
  Journal Article
• Place as Subject:
  California ; OSHA Region 9
• CIO:
  National Institute for Occupational Safety and Health (NIOSH)
• Topic:
  Workplace Safety & Health
• Location:
  North America
• Pages in Document:
  347-360
• Volume:
  87
• Issue:
  1
• NIOSHTIC Number:
  nn:20032377
• Citation:
  Chem-Biol Interact 1993 Jun; 87(1-3):347-360
• Contact Point Address:
  Thomas C. Thomas, Brigham and Women's Hospital, Thorn Building, Room 1123, 75 Francis St., Boston, MA 02115, USA
• Federal Fiscal Year:
  1993
• Performing Organization:
  University of California - Davis
• Peer Reviewed:
  True
• Start Date:
  19900930
• Source Full Name:
  Chemico-Biological Interactions
• End Date:
  20020929
• Collection(s):
  National Institute for Occupational Safety and Health
• Main Document Checksum:
  urn:sha-512:396d4af39810acd20523fd2559154f26dc7872282f64282e3b34f091bfd60f90dcdccd8445fb3274e109501cb4142663e46180361b0fa851b857b0391eb1f411
• Download URL:
  https://stacks.cdc.gov/view/cdc/186383/cdc_186383_DS1.pdf
• File Type:
  [PDF - 956.12 KB ]