A new type of O2--generating tool for oxidative stress studies by remodeling neutrophil NADPH oxidase

Minoru Tamura, Teruaki Nagasawa, Tomoyuki Tange, Kei Miyano, Shin Ichi Kobayashi, Toshiki Nakai, Satoshi Miura

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

The effects of reactive oxygen species on cells have attracted much attention in relation to redox regulation and oxidative stress-related diseases. Superoxide (O2-) is the reactive oxygen species primarily formed in biological systems. However, no convenient O2 --generating device has been available for use in cell or tissue culture. The neutrophil NADPH oxidase, a professional enzyme for killing bacteria, has a high ability to produce O2-. However, the cell-free activation process requires several protein factors and an anionic amphiphile, and moreover, the activation is transient. To utilize the enzyme as an O2- generator, we improved the cell-free activation method by remodeling regulatory components, optimizing lipid composition, and modifying the mixing conditions. We established a new method to produce an active enzyme that is stable, efficient, and preservable. As an application, we examined the effect of the device on cultured HEK293 cells and observed that it caused cell death. This system has several advantages over the xanthine oxidase system often used. The new device will be useful for studies of oxidative stress and related diseases.

Original languageEnglish
Pages (from-to)421-429
Number of pages9
JournalJournal of Biotechnology
Volume120
Issue number4
DOIs
Publication statusPublished - Dec 6 2005

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology

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