TEMPERATURE ACTIVATION OF THE UREASE-UREA SYSTEM USING CRUDE AND CRYSTALLINE UREASE.
1. The hydrolysis of urea catalyzed by jack bean meal has been followed by determining colorimetrically after Nesslerization the ammonia nitrogen, and volumetrically the carbon dioxide liberated at successive intervals during the reaction. During the early part of hydrolysis the rate of ammonia or carbon dioxide liberation is constant for all the urease solutions which were used. 2. When log rate of NH(3) or CO(2) formation was plotted against 1/T, the points fell along a straight line, the slope of which corresponded to an activation energy of either 8,700 or 11,700 calories per gram mol. Frequently urease, when dissolved in sulfite solution, was characterized by an activation energy of 11,700 below and 8,700 above the critical temperature of about 23 degrees C. At high temperatures the plotted points fell off from the curve due to temperature inactivation. 3. Essentially the same results on temperature activation were obtained with crude jack bean meal, Arlco urease, crystalline urease not recrystallized, and crystalline urease once recrystallized. The temperature characteristic which was obtained depended in part upon the composition of the medium. When dissolved in water, or aqueous solutions of glycerine, KCN, Na(2)S(2)O(2), cystine, Na(2)SO(4), and K(4)Fe(CN)(6), the temperature characteristic or micro of urease is 8,700. On the other hand, when urease is dissolved in solutions of K(3)Fe(CN)(6) or H(2)O(2) the micro value is 11,700. When dissolved in a solution containing Na(2)SO(3) and NaHSO(3) the micro value may be either 8,700 or 11,700 over the whole temperature range, or 11,700 below and 8,700 above 23 degrees C. 4. When crystalline urease is dissolved in varying mixtures of K(4)Fe(CN)(6) and K(3)Fe(CN)(6), the temperature characteristic depends upon the oxidation-reduction potential of the digest. When E(h) is greater than +0.46 volt micro = 11,700, when less than +0.42 volt micro = 8,700, when between +0.42 - +0.46 micro = 11,700 below and 8,700 above the critical temperature. 5. It is suggested that in reducing or in indifferent solutions the configuration of the urease molecule (as determined especially by SH groups present) is such that the activation energy is 8,700 calories. In oxidizing solutions the urease molecule has been so altered (perhaps by the oxidation of the SH groups) as to be partly inactivated and now has an activation energy of 11,700. Such changes in the urease molecule are reversible (unless oxidation has proceeded too far) and are accompanied by a corresponding change in the activation energy.
Sizer
I W
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Laboratory of Physiology and Biochemistry, Massachusetts Institute of Technology, Cambridge.