Evidence for conformers of rabbit muscle adenylate kinase.
Changes of the apparent Mr values and the circular dichroism patterns suggest the existence of three relatively stable conformers of rabbit muscle adenylate kinase (RMAK). The effects of dithiothreitol (DTT) which stimulates activity, pH, the substrates, and ATP on the Mr value and the Stokes radius of RMAK were determined from gel filtration data, and apparent Mr values near 22,000, 26,000, and 29,000 resulted. Substrates generated multiple Mr values, suggesting the presence of multiple conformers of RMAK. The higher apparent Mr values were obtained in the presence of DTT and at the higher substrate concentrations, indicating more open conformations. The effect of the substrates on the conformation of RMAK is discussed in relation to the kinetic mechanism of this random bireactant system. Circular dichroism studies were undertaken in order to observe any changes in the secondary structures of RMAK in relation to changes of the Mr values. The secondary structure composition of RMAK, determined under our conditions, does not agree with results determined from crystallographic studies. The gel filtration and the CD studies suggest that above pH 7 a more open conformation of RMAK obtains in the presence of DTT. The results of these studies are discussed with reference to the location of the active sites.
Russell
P J
PJ
Department of Biology and Chemistry, University of California, San Diego, La Jolla 92023.
Chinn
E
E
Williams
A
A
David-Dimarino
C
C
Taulane
J P
JP
Lopez
R
R
D000263:Adenylate Kinase / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D002942:Circular Dichroism; D004229:Dithiothreitol / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008970:Molecular Weight; D009132:Muscles / Q000201:enzymology*; D010770:Phosphotransferases / Q000378:metabolism*; D011487:Protein Conformation; D011817:Rabbits
D007527:Isoenzymes; D010770:Phosphotransferases; D000263:Adenylate Kinase; D004229:Dithiothreitol