PLANT PHYSIOLOGY , Vol 103, Issue 3 733-739, Copyright © 1993 by American Society of Plant Biologists
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METABOLISM AND ENZYMOLOGY |
Purification and Properties of a Monofunctional Imidazoleglycerol-Phosphate Dehydratase from Wheat
J. Mano, M. Hatano, S. Koizumi, S. Tada, M. Hashimoto and A. Scheidegger
International Research Laboratories, Ciba-Geigy (Japan) Ltd., P.O. Box 1, Takarazuka, Hyogo 665, Japan
Imidazoleglycerol-phosphate dehydratase (EC 4.2.1.19) activity was detected
in extracts of several monocotyledonous and dicotyledonous plants using a
newly developed assay method. The enzyme was purified 114,000-fold (to
apparent homogeneity) from wheat germ by five chromatographic steps. Its
native relative molecular weight (Mr) was determined to be 600,000 to
670,000, and it consists of identical subunits of Mr 25,500. In wheat germ,
the dehydratase, unlike those of prokaryotic origin, is not associated with
histidinol phosphatase activity. The reaction product was identified as
imidazoleacetol phosphate (IAP) by comparing it with synthetic IAP as an
authentic reference. The Km value for imidazoleglycerol phosphate was 0.36
mM at the optimal pH of 6.6. The enzyme required a reducing agent, such as
2-mercaptoethanol or dithiothreitol, and Mn2+ for maximal activity.
3-Amino-1,2,4-triazole competitively inhibited the activity with a Ki value
of 46 [mu]M. The purification of imidazoleglycerol-phosphate dehydratase
from wheat germ and histidinol dehydrogenase from cabbage (A. Nagai, A.
Scheidegger [1991] Arch Biochem Biophys 284: 127-132) suggests that at
least the second half of the histidine biosynthesis in plants is identical
to that in microorganisms.
