Studies on the effect of temperature on the activity and stability of cyanobacterial ADP-glucose pyrophosphorylase.
The effect of temperature on the activity and stability of ADPglucose pyrophosphorylase from Anabaena PCC 7120 was studied. Experimental optima temperatures were found around 37-40 degrees C or 42-45 degrees C, depending on the absence or the presence of allosteric effectors in the assay medium, respectively. In the range of temperature where the enzyme is stable, curved Arrhenius plots were obtained, indicating a transition temperature between 9 and 12 degrees C. Since these results were observed for both the forward and reverse reaction, with two different sets of substrates and two entirely different assay procedures, it seems unlikely that the effect can be on any component of the system other than the enzyme itself. Results suggest that cyanobacterial ADPglucose pyrophosphorylase undergoes conformational changes at different temperatures, rendering structures with different catalytic efficiencies. The different structures of the enzyme were visualized by emission fluorescence. ADPglucose pyrophosphorylase was irreversibly inactivated when exposed to temperatures above 40 degrees C. Inactivation was dependent on temperature and followed first order kinetics. The substrate, ATP, and the allosteric effectors, 3PGA and Pi, effectively protected the enzyme against thermal inactivation. Protection afforded by ATP was affected by MgCl2. These results suggest that the binding of the effectors to the enzyme resulted in conformational changes of the protein, rendering structures more stable to temperature treatments. Similar structures could be adopted by the enzyme in different environments, since the higher stability was observed in media containing either high ionic strength or high hydrophobicity.
Gómez-Casati
D F
DF
Instituto Technológico de Chascomús (IIB-INTECH, CONICET), Argentina.
Preiss
J
J
Iglesias
A A
AA
D000245:Adenosine Diphosphate Glucose / Q000096:biosynthesis; D000255:Adenosine Triphosphate / Q000378:metabolism; D000494:Allosteric Regulation; D017033:Anabaena / Q000201:enzymology*; D004795:Enzyme Stability; D051599:Glucose-1-Phosphate Adenylyltransferase; D007700:Kinetics; D009713:Nucleotidyltransferases / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013696:Temperature
D000245:Adenosine Diphosphate Glucose; D000255:Adenosine Triphosphate; D009713:Nucleotidyltransferases; D051599:Glucose-1-Phosphate Adenylyltransferase