Fusicoccin action in cell-suspension cultures of Corydalis sempervirens Pers. |
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Authors: | Simone Schulz Edith Oelgemöller Elmar W Weiler |
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Institution: | (1) Lehrstuhl für Pflanzenphysiologie, Ruhr-Universität, Postfach 102148, W-4630 Bochum, Germany |
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Abstract: | Mid-log-phase cell suspensions of Corydalis sempervirens Pers., when incubated in micromolar or submicromolar concentrations of fusicoccin, strongly acidified the culture medium. High-affinity fusicoccin-binding sites were found in microsomes prepared from these cells using the radioligand 3H]-9 -norfusicoccin-8 -alcohol. Binding was saturable with an apparent dissociation constant (K
d) of 2.8 nM, a pH optimum of 6.0, a temperature optimum of 35° C and was rapid (t1/2 = 8 min). The site abundance was 0.76±0.17 pmol · (mg of protein)–1. In the same membrane preparations, the K+, Mg2+-ATPase (EC 3.6.1.3) was characterized. The enzyme was highly vanadate-sensitive (IC50=6.5 M) and nucleotide-specific (ATP NTP), had a pH optimum of 6.2, an apparent K
m for ATP of 0.23±0.12 mM, and V
max of 10.6±1.8 nkat (mg of protein)–1. Fusicoccin doubled V
max and lowered, by a factor of 2, the apparent K
m for ATP of the enzyme when the cells were incubated with the toxin for 30 min prior to homogenization of the cells. The stimulation of the enzyme was also pronounced when fusicoccin was added to the homogenization medium just prior to homogenization of the cells, but was slight to zero when the toxin was added at the microsomal stage. The pronounced stimulatory effect of fusicoccin on the ATPase was seen at pH 7.1, i.e. at a pH typical for the cytoplasmic compartment, but was not detectable at pH 6.2, the pH optimum of the enzyme. The implications of these findings for an understanding of fusicoccin action are discussed.Abbreviations 3H]ABE-FC
9 -nor-8 -(4-azido-3,5-3H]-benzoyl-diaminoethyl)-fusicoccin
- FC
fusicoccin
- FCol
9 -norfusicoc-cin-8 -alcohol
- Mes
2(N-morpholino)ethanesulfonic acid
This work was supported by the Deutsche Forschungsgemeinschaft, Bonn, FRG and the Fonds der Chemischen Industrie, Frankfurt, FRG (literature provision). |
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Keywords: | Cell culture (fusicoccin binding) Corydalis Fusicoccinbinding protein K+ Mg2+ ATPase Proton extrusion |
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