Selective Inactivation of an Extra-cytoplasmic Acid Phosphatase of Yeast-like Cells of Sporothrix schenckii by Sodium Fluoride

Abstract

Suspensions of intact, yeast-like cells of Sporothrix schenckii exhibited an acid phosphatase (EC 3 . 1 .3.2) activity against pnitrophenyl phosphate of about 5 IU (g dry wt)-l, without recourse to membrane perturbation. This extra-cytoplasmic acid phosphatase was reversibly and competitively inhibited by orthophosphate (Ki = 2 mM at pH 5) but unaffected by L( +)-tartrate (in contradistinction to some of the cytoplasmic acid phosphatases of the same organism). Inactivation by NaF of the extra-cytoplasmic isoenzyme was irreversible and followed first order kinetics; sensitivity to NaF was decreased by the presence of citrate, phosphate or substrate. Neither K , (0.3 mM at pH 5) nor V,,, for this enzyme in acetate buffer was greatly affected by pH in the range 3-5 but the first order rate constant for inactivation by NaF was strongly dependent on pH (maximum at pH 3.5). Crude cell-free extracts of yeast cells had nine electrophoretically distinct acid phosphatase activity bands and, on the basis of the pattern of inhibitors, the extra-cytoplasmic activity was identified as Y-I, an isoenzyme that barely penetrates standard polyacrylamide gel electropherograms. Additional evidence for the assignment came from selective inactivation of this isoenzyme by short treatments of intact cells with NaF under conditions that did not allow penetration of the plasma membrane by the inhibitor and did not kill the cells.

Acid phosphatase specific to arbuscular mycorrhizal infection in marigold and possible role in symbiosis

“…fluoride was only effective in acidic zone. This finding corroborates the observation of Arnold et al (1987).…”

“…The adsorption to lectin (Con A) and the subsequent elution with a-methyl-o-glucoside suggested that the enzyme might be a glycoprotein as reported for other acid phosphatases (Van Etten and Saini 1977;Bhargava and Sachar 1987). Citrate (10 or 100 mM) did not prevent the inhibitory effect of fluoride in contrast to the yeast acid phosphatase (Arnold et al 1987). Phosphonic acid (such as ABP), an analogous compound of phosphate ester, has been shown to be an inhibitor of non-specific acid and alkaline phosphatases and used in affinity chromatography (Landt et al 1978;Dissing et al 1979).…”

“…3.1.3.2) (Uehara et al 1974;Hasegawa et al 1976;Panara et al 1990). The inhibition of the ISPase activity by fluoride, molybdate, vanadate, and phosphate, and the lack of i nhi bitory effect of chelating agents, such as EDT A, are typical characteristics of acid phosphatase (Uehara et al 1974;Hasegawa et al 1976;Ninomiya et al 1977;Arnold et al 1987;Duff et al 1989). Especially, the effective cleavage of the pyrophosphate bond suggests that the enzyme may be an acid phosphatase from plant (Uehara et al 1974;Hasegawa et al 1976;Ninomiya et al 1977;Villareal and Juliano 1977;Szabo-Nagy et al 1987;Panara et al 1990).…”

Regulators of GTP‐binding proteins cause morphological changes in the vacuole system of the filamentous fungus, Pisolithus tinctorius

“…The results from our controls show that NaF itself can be a powerful stimulant of tubule formation, and the rise in tubule frequency when the concentration of NaF was raised from 5 to 20 mM mirrored that seen when com- paring the response to the 5 mM NaF/30 M AlCl 3 and 20 mM NaF/10 M AlCl 3 combinations. We do not know the reason for the response to NaF, but this compound is well known as an inhibitor of many phosphatases, kinases, and ATPases in fungi and other organisms [e.g., Arnold et al, 1987;Pinkse et al, 1999;Vargas et al, 1999], which could directly or indirectly influence tubule-forming activity. It should be noted here that some recent studies reporting the effects of aluminium fluoride on processes possibly regulated by GTP-binding proteins have not checked for non-specific effects of fluorides.…”

Comparison of phosphatase localization in the intraradical hyphae of arbuscular mycorrhizal fungi, Glomus spp. and Gigaspora spp.

“…At that time, the root cortex was torn with needles to expose the fungal tissue to the staining solution. The effects of 10 mM NaF (acid phosphatase inhibitor (Arnold et al 1987)), 5 mM EDTA and 10 mM KCN (both are alkaline phosphatase inhibitors (Belle, 1972)) on the phosphatase activity in the hyphae were also examined. Acid and alkaline phosphatase activities in the fungal tissue were identified by the azo dye method (Tisserant et al, 1993).…”