Scripps VIVO scripps research logo

  • Index
  • Log in
  • Home
  • People
  • Organizations
  • Research
  • Events
Search form
As of April 1st VIVO Scientific Profiles will no longer updated for faculty, and the link to VIVO will be removed from the library website. Faculty profile pages will continue to be updated via Interfolio. VIVO will continue being used behind the scenes to update graduate student profiles. Please contact helplib@scripps.edu if you have questions.
How to download citations from VIVO | Alternative profile options

Expression and immunoaffinity purification of human inducible nitric-oxide synthase - inhibition studies with 2-amino-5,6-dihydro-4H-1,3-thiazine

Academic Article
uri icon
  • Overview
  • Identity
  • Additional Document Info
  • View All
scroll to property group menus

Overview

authors

  • Calaycay, J. R.
  • Kelly, T. M.
  • MacNaul, K. L.
  • McCauley, E. D.
  • Qi, H. B.
  • Grant, S. K.
  • Griffin, Patrick
  • Klatt, T.
  • Raju, S. M.
  • Nussler, A. K.
  • Shah, S.
  • Weidner, J. R.
  • Williams, H. R.
  • Wolfe, G. C.
  • Geller, D. A.
  • Billiar, T. R.
  • MacCoss, M.
  • Mumford, R. A.
  • Tocci, M. J.
  • Schmidt, J. A.
  • Wong, K. K.
  • Hutchinson, N. I.

publication date

  • November 1996

journal

  • Journal of Biological Chemistry  Journal

abstract

  • Recombinant human inducible nitric-oxide synthase (rH-iNOS) was expressed in the baculovirus system and purified by a novel immunoaffinity column. rH-iNOS and its native counterpart from cytokine-stimulated primary hepatocytes exhibited similar molecular mass of 130 kDa on SDS-polyacrylamide gel electrophoresis, recognition by antipeptide antibodies, specific activities, and IC50 values for inhibitors. The active dimeric form exhibited a specific activity range of 114-260 nmol/min/mg at 37 degrees C and contained 1.15 +/- 0.04 mol of calmodulin/monomer. The enzyme exhibited a Soret lambdamax at 396 nm with a shoulder at 460 nm and contained 0. 28-0.64 mol of heme/monomer. Dithionite reduction under CO yielded an absorbance maximum at 446 nm, indicating a P450-type heme. Imidazole induced a type II difference spectrum, reversible by L-Arg. 2-Amino-5,6-dihydro-4H-1,3-thiazine (ADT) was competitive versus L-Arg (Ki = 22.6 +/- 1.9 nM), reversed the type II difference spectrum induced by imidazole (Kd = 17.7 nM), and altered the CO-ferrous absorbance of rH-iNOS. L-Arg did not perturb the CO-ferrous adduct directly, but it partially reversed the ADT-induced absorbance shift, indicating that both bind similarly to the protein but interact differently with the heme.

subject areas

  • Chromatography, High Pressure Liquid
  • Enzyme Induction
  • Humans
  • Kinetics
  • NADP
  • Nitric Oxide Synthase
  • Radiation-Protective Agents
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
  • Thiazines
scroll to property group menus

Identity

International Standard Serial Number (ISSN)

  • 0021-9258

PubMed ID

  • 8910438
scroll to property group menus

Additional Document Info

start page

  • 28212

end page

  • 28219

volume

  • 271

issue

  • 45

©2022 The Scripps Research Institute | Terms of Use | Powered by VIVO

  • About
  • Contact Us
  • Support