In vitro activation of the NADPH oxidase by fluoride. Possible involvement of a factor activating GTP hydrolysis on Rac (Rac-GAP).
The possible mechanism of activation of the NADPH oxidase by fluoride was investigated in the cell-free system. It is shown that the stimulatory effect of fluoride is inhibited by guanosine 5'-O-(2-thiodiphosphate) (GDP[S]) and potentiated by GTP. The effect of fluoride is not additive with GTP[S]. Fluoride activation requires the presence of Mg2+ in millimolar concentration but is independent of Al3+. The activating effect of fluoride is preserved in solubilized membrane extract after removal of the majority of heterotrimeric GTP-binding proteins by immunoadsorption. Fluoride has no direct action either on the nucleotide exchange of GTP hydrolysis of the isolated Rac protein. In contrast, fluoride effectively inhibits Rac-GTPase activity enhanced by a membrane component. In this way, fluoride could prolong the prevalence of Rac in the GTP-bound state and, as a consequence, activate NADPH oxidase. The possibility of the involvement of a membrane-bound Rac GTPase-activating protein activity in the physiological regulation of the enzyme is raised.
Wölfl
J
J
Department of Physiology, Semmelweis Medical University, Budapest, Hungary.
Dagher
M C
MC
Fuchs
A
A
Geiszt
M
M
Ligeti
E
E
D000818:Animals; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism; D020690:GTPase-Activating Proteins; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006153:Guanosine Diphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000378:metabolism; D006868:Hydrolysis; D007700:Kinetics; D009247:NADH, NADPH Oxidoreductases / Q000097:blood*; D019255:NADPH Oxidases; D009504:Neutrophils / Q000378:metabolism; D011506:Proteins / Q000378:metabolism*; D012969:Sodium Fluoride / Q000494:pharmacology*; D013481:Superoxides / Q000097:blood; D013552:Swine; D013873:Thionucleotides / Q000494:pharmacology; D013997:Time Factors
D020690:GTPase-Activating Proteins; D011506:Proteins; D013873:Thionucleotides; D013481:Superoxides; D006153:Guanosine Diphosphate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); C023427:guanosine 5'-O-(2-thiodiphosphate); D006160:Guanosine Triphosphate; D012969:Sodium Fluoride; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases; D019204:GTP-Binding Proteins