ATP-GTP-BINDING PROTEINS AND ENDOGENOUS ADP-RIBOSYL TRANSFERASE IN Lemna paucicostata 441

Abstract— A crude extract containing membrane components of Lemna paucicostata was treated with 1% Lubrol PX and fractionated by gel nitration. Binding activities to non-hydrolyzable analogues of ATP, ³⁵S]ATPγS (adenosine 5';γ-thio]triphosphate) and that of GTP, ³⁵S]GTPγS (guanosine 5'γ-thiojtriphosphate) were detected in some fractions, and these activities were prevented in the presence of 0.1 mM ATP or GTP. ATP and GTP were 2 to 3 orders of magnitude more effective than CTP or UTP in preventing this binding activity. These fractions showed ATPase and GTPase activities with 1 nM γ-³²P]ATP or γ–³²P]GTP substrate. Analyses by sodium dodecyl sulfate polyacrylamide gel electrophoresis of these fractions after binding with ³⁵S]ATPγS or ³⁵S]GTP-γ S revealed that these fractions contained ³⁵S]ATPγS and ³⁵S]GTPγS binding proteins with molecular weights of 53 000 and 60 000, respectively. Both of these proteins were ³²P]ADP-ribosylated by endogenous ADP-ribosyl transferase. Three proteins with molecular weights of 11 000, 12 000 and 13 000 which could bind ³⁵S]ATP7S or -³⁵S]GTP-γ S were ADP-ribosylated by endogenous ADP-ribosyl transferase. Pertussis toxin stimulated ADP-ribosylation of these proteins. Four proteins with molecular weight of 37 000, 50 000, 80 000 and 115 000 with P^SS]ATP7S and ^,3S]GTP7S binding activities were also detected. The signal transduction of light to underlying clock mechanism in Lemna may be controlled by ATP-GTP-binding proteins and by the ADP-ribosylation of these proteins.