Enzymatic characterization of recombinant murine inducible nitric-oxide synthase

Moss, David W., Wei, Xiao-Qing ORCID: https://orcid.org/0000-0002-6274-8503, Liew, Foo Y., Moncada, Salvador and Charles, Ian G. 1995. Enzymatic characterization of recombinant murine inducible nitric-oxide synthase. European Journal Of Pharmacology-Molecular Pharmacology Section 289 (1) , pp. 41-48. 10.1016/0922-4106(95)90166-3

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Abstract

A complementary DNA (cDNA) encoding murineinduciblenitricoxidesynthase was cloned from activated J774 macrophages. Expression of this cDNA in a baculovirus-insect cell system allowed comparison of the recombinant enzyme with the native homologue. Western blot analysis of activated J774 and baculovirus-infected insect cell cytosols demonstrated reactivity against a protein of 135 kDa. Kinetic studies on the recombinant and native enzymes revealed an absolute requirement for L-arginine and NADPH in order to achieve full activity. In addition, both enzymes were found to have similar maximum velocities and Km values for these two substrates. The nitricoxidesynthase antagonists N-guanidino monomethyl L-arginine and N-iminoethyl L-ornithine inhibited both enzymes at a similar rate. Furthermore, comparable concentrations of inhibitor were required to achieve half maximal enzyme inhibition. These results indicate that recombinantinducible NO synthase appears to be pharmacologically indistinguishable from the native enzyme.

Item Type:	Article
Date Type:	Publication
Status:	Published
Schools:	Dentistry
Subjects:	R Medicine > RK Dentistry
Uncontrolled Keywords:	Nitricoxide(NO); Nitricoxidesynthase; Cloning; Expression; Baculovirus; Endotoxic shock
Publisher:	Elsevier
Last Modified:	21 Oct 2022 09:13
URI:	https://orca.cardiff.ac.uk/id/eprint/35639

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