Influence of Growth Temperature on Structure, Thermostability and Kinetic Properties of the ATPase Coupling Factor AF1 of a Thermophilic Blue‐Green Alga (Cyanobacterium)

Abstract

The coupling factor AF1 isolated from cells of the thermophilic blue-green alga M. laminosus grown at 40.degree. C, 50.degree. C and 60.degree. C was investigated and compared with the chloroplast coupling factor CF1. The structure of AF1 is affected by the growth temperature. The AF1 from 60.degree. C shows a split .alpha.-band in dodecylsulfate/polyacrylamide gel electrophoresis which is not observed in AF1 from 40.degree. C or from 50.degree. C. Only the AF1 from 60.degree. C aggregates with allophycocyanin. The AF1 from 60.degree. C is stable up to 85.degree. C for 60 min whereas the AF1 from 40.degree. C and 50.degree. C denature between 65.degree. C and 70.degree. C. CF1 is much less stable. In contrast to the structure, the kinetic properties of the coupling factor are not influenced by the growth temperature. The reaction rates of all 3 AF1 preparations have the same temperature optimum which is also identical to that of CF1. The kinetic properties of the AF1 from 50.degree. C were determined in more detail. In the absence of ADP the saturation curve for ATP shows Michaelis-Menten kinetics with Km = 480 .mu.M. With the addition of ADP this curve becomes sigmoidal and the reaction rate decreases. This behavior and the Hill values suggest an allosteric enzyme with 2 active sites acting cooperatively. ADP as a product of the ATPase reaction inhibits the enzyme. The inhibition is not simply competitive, it also influences the cooperativity of the active sites.