J. J. Robertson, H. Orin Halvorson
1957 Journal of Bacteriology  
The term maltozymase designates the enzyme system involved in the production of carbon dioxide from maltose in yeast. The discovery of a hydrolytic a-glucosidase led Fisher (1895) to propose that the following reactions represent the action of maltozymase: maltose ki > glucose k2 4 C02, etc. (1) Recent studies on the induced synthesis of a-glucosidase (Halvorson and Spiegelman, 1957) support Fisher's hypothesis. During the early stages of induction, the rate of the a-glucosidase reaction (k1)
more » ... mits the over-all activity of maltozymase. However, the observation that maltose is not metabolized, while glucose is, under conditions in which a-glucosidase would be expected to operate in vivo (Willstatter and Bamann, 1926; Leibowitz and Hestrin, 1945) suggests that maltozymase may contain other enzyme reactions. Since the reactions produced by maltozymase include all those involved in glucose metabolism, it is clear that the appropriate method for searching for other components of maltozymase is to establish conditions in which its activity can be varied independently from glucose metabolism. Such conditions are provided when yeast cells grown in maltose are incubated aerobically with glucose in nitrogen-free medium (Spiegelman and Reiner, 1947) . The resulting loss of maltozymase activity has been termed deadaptation. Although the phenomenon of deadaptation has led to speculation about the existence of competitive reac-I tions between different enzyme-forming systems (Spiegelman and Dunn, 1947) , the basis of the loss of maltozymase activity is not understood. Dinitrophenol and azide prevent deadaptation (Spiegelman and Reiner, 1947) , suggesting that the process requires energy. Deadaptation is prevented also by nitrogen or by the inducing agent. This paper reports the results of a study undertaken to determine the various components of the maltozymase system. The identity of these and their behavior during deadaptation are described. METHODS AND MATERIALS A diploid representative of Saccharomyces cerevisiae strain LK2G12, was used. Cells were grown in stationary flasks at 30 C in a medium prepared by adding the following to 1 L of water: peptone (Difco), 5 g; yeast extract (Difco), 2.5 g; (NH4)2SO4, 2 g; CaCl2, 0.25 g; MgSO4, 0.25 g; KH2PO4, 2 g; and glucose (or maltose), 40 g. Cells in the log phase (12 hr) were harvested by centrifugation, washed twice in cold water, and resuspended, either in water or in a buffer free of nitrogen and carbohydrate (Halvorson and Spiegelman, 1952) , to a density of 2.84 mg dry weight of yeast per ml. To induce the enzyme synthesis, cells were suspended in buffer supplemented with 3 per cent maltose. In the deadaptation experiments, glucose was substituted for maltose as the energy source. The suspensions were incubated aerobically at 30 C, either in a standard Warburg apparatus or in Erlenmeyer flasks, on a rotary platform shaker. Maltozymase was followed in the washed intact cells by the two-cup method, after stabilization by ultraviolet light (Halvorson and Spiegelman, 1954) . Three methods were employed to rupture the cells: (1) Fast-dried preparations were obtained by lyophilizing samples which had previously been shell-frozen at -30 C. After evacuation for 4 hr, the samples were stored overnight over 186 on May 9, 2020 by guest
doi:10.1128/jb.73.2.186-198.1957 fatcat:7rqa7vjfovde7fom6agkeq3rti