PROSTHETIC GROUPS, COENZYMES AND ENZYMES 



hydrochloric acid. The value of ki increases fourfold from 

 13.5 to 23.5° C. 



THE ASSOCIATION VELOCITY (27) 



In order to study the association velocity constant, ki, as 

 a function of /?H we used acetate from p\l 4 to 7, and glycine 

 from /?H 7 to 10.5, since neither of these buffer substances was 

 found to interfere with the "on" velocity. The value of ki was 

 found to vary with /?H in a most interesting way (see Figure 3) . 

 From/?H 4 to 8 A;i at 23.5 ° C. followed a monovalent dissociation 

 curve with j&K' = 6. This coincides with the second dissocia- 

 tion constant of the phosphoric acid residue of FMN. From 

 pH. 8 to 9 there is a new increase in kx up to a maximum around 

 pYi 9 at 23.5 ° G. or 9.3 at 13.5 ° C. With increasing alkalinity 



TABLE II 



The Inhibition of ki by Polyvalent Anions and Its Counteraction by Mono- 

 valent Anions {pU. = 7.0, 23.5° G.) 



ki falls off very rapidly; ^i is = 1.5 X 10^ M~'^ sec.~^ at />H 9, 

 thus roughly half as great as ki for DPNH + ADH, and five 

 times greater than the "on" velocity for DPN + ADH {cf. 

 below). Anions decrease ki\ the higher the charge, the stronger 

 the effect. Secondary phosphate, for example, inhibited ki 

 very considerably, whereas primary phosphate did not. The 

 inhibitory effect of polyvalent anions was to a great extent 

 counteracted by monovalent anions. 



287 



