Characterization of an ecto-ATPase activity in Fonsecaea pedrosoi.
In this work, we characterized an ecto-ATPase activity in intact mycelial forms of Fonsecaea pedrosoi, the primary causative agent of chromoblastomycosis. In the presence of 1 mM EDTA, fungal cells hydrolyzed adenosine-5'-triphosphate (ATP) at a rate of 84.6 +/- 11.3 nmol Pi h(-1) mg(-1) mycelial dry weight. The ecto-ATPase activity was increased at about five times (498.3 +/- 27.6 nmol Pi h(-1) mg(-1)) in the presence of 5 mM MgCl2, with values of Vmax and apparent Km for Mg-ATP(2-) corresponding to 541.9 +/- 48.6 nmol Pi h(-1) mg(-1) cellular dry weight and 1.9 +/- 0.2 mM, respectively. The Mg2+-stimulated ecto-ATPase activity was insensitive to inhibitors of intracellular ATPases such as vanadate (P-ATPases), bafilomycin A1(V-ATPases), and oligomycin (F-ATPases). Inhibitors of acid phosphatases (molybdate, vanadate, and fluoride) or alkaline phosphatases (levamizole) had no effect on the ecto-ATPase activity. The surface of the Mg2+ -stimulated ATPase in F. pedrosoi was confirmed by assays in which 4,4'-diisothiocyanostylbene-2,2'-disulfonic acid (DIDS), a membrane impermeant inhibitor, and suramin, an inhibitor of ecto-ATPase and antagonist of P2 purinoreceptors. Based on the differential expression of ecto-ATPases in the different morphological stages of F. pedrosoi, the putative role of this enzyme in fungal biology is discussed.
Collopy-Junior
Italo
I
Instituto de Bioquímica Médica, Universidade Federal do Rio de Janeiro, CCS, Bloco H, Cidade Universitária, Ilha do Fundão, 21541-590, Rio de Janeiro, Brazil.
Kneipp
Lucimar F
LF
da Silva
Fernanda C
FC
Rodrigues
Marcio L
ML
Alviano
Celuta S
CS
Meyer-Fernandes
José Roberto
JR
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D001203:Ascomycota / Q000166:cytology / Q000201:enzymology* / Q000502:physiology; D003067:Coenzymes / Q000494:pharmacology; D004492:Edetic Acid / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005459:Fluorides / Q000494:pharmacology; D007978:Levamisole / Q000494:pharmacology; D018942:Macrolides / Q000494:pharmacology; D015636:Magnesium Chloride / Q000494:pharmacology; D009840:Oligomycins / Q000494:pharmacology; D013172:Spores, Fungal / Q000201:enzymology; D014638:Vanadates / Q000494:pharmacology
D003067:Coenzymes; D004791:Enzyme Inhibitors; D018942:Macrolides; D009840:Oligomycins; D015636:Magnesium Chloride; C057620:bafilomycin A; D007978:Levamisole; D014638:Vanadates; D000255:Adenosine Triphosphate; D004492:Edetic Acid; D000251:Adenosine Triphosphatases; D005459:Fluorides