[25 Redox Sensor Function of Metallothioneins

By james P. fabisiak, gregory g. borisenko, shang-xl llu, Vladimir A. Tyurin, Bruce R. Pitt, and Valerian E. Kagan

Introduction

Metallothioneins (MTs) are low molecular weight (approximately 6000) cysteine-rich (30%) metal-binding proteins.1 Originally discovered and isolated as a cadmium-binding protein from horse kidney, it is now apparent that the multi-isoform family of MT proteins can serve to protect cells and animals from the toxic effects of metals,2 reactive electrophiles,3 and reactive oxygen/nitrogen species (ROS/RNS).4-5

The 20 cysteines contained in MTs are highly conserved across species; are arranged into two distinct thiolate clusters, termed a and fi: and are fundamental to the metal-binding function through the formation of sulfur-metal bonds.6 For metal binding to occur, the cysteine SH groups must be in their reduced state. Copper binding to MT has been shown to be directly proportional to the content of reduced SH groups7 and oxidative or nitrosative stress can, in fact, promote copper

1 J. H. R. Kagi and A. Schaffer, Biochemistry 27, 8509 (1988).

2 M. P. Waalkes and P. L. Goering, Chem. Res. Toxicol. 3, 281 (1990).

3 J. S. Lazo and B. R. Pitt, Annu. Rev. Pharmacol. Toxicol. 35, 635 (1995).

4 B. R. Pitt, M. Schwarz, E. S. Woo, E. Yee, K. Wasserloos, S. Tran, W. Weng, R. J. Mannix, S. A. Watkins, Y. Y. Tyurina, V. Tyurin, V. E. Kagan, and J. S. Lazo, Am. J. Physiol. (Lung Cell. Mol. Physiol.) 273, L856(1997).

5 M. A. Schwarz, J. S. Lazo, J. c. Yalowich, I. Reynolds, V. E. Kagan, V. Tyurin, Y.-M. Kim, S. Watkins, and B. R. Pitt, J. Biol. Chem. 269, 15238 (1994).

6 J. D. Otvos and I. M. Armitage, Proc. Natl. Acad. Sci. U.S.A. 77, 7094 (1985).

7 J. P. Fabisiak, V. A. Tyurin, Y. Y. Tyurina, G. G. Borisenko, A. Korotaeva, B. R. Pitt, J. S. Lazo, and V. E. Kagan, Arch. Biochem. Biophys. 363, 171 (1999).

Conclusion

ROS regulate a variety of vascular signaling pathways. Using an ELISA, we have shown that superoxide regulates ECE activity. Indirect measurements and direct measurements show that superoxide inhibits ECE by ejecting zinc from the enzyme. Several pathways in the cardiovascular system involve the activity of converting enzymes, including the renin-angiotensin-aldosterone pathway. Techniques that measure protease activity, and measure the effect of ROS on proteases, may reveal novel mechanisms by which radicals regulate cardiovascular signaling.

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