272 VINCENT G. ALLFREY 



It remains to be seen whether some transient derivative of alanine is formed 

 in the course of this accumulation process. 



Competition experiments using non-radioactive D-alanine and l- 

 alanine before adding DL-[i-^^C]-alanine have shown that only the naturally 

 occurring L-isomer of the amino acid competes with radioactive alanine for 

 transport. It follows that the D-isomer of the amino acid is not actively 

 accumulated by the nucleus [29, 46]. The stereo-specificity of the reaction 

 suggests that amino acid transport into the nucleus involves the operation 

 of an enzymic mechanism. 



The alanine transport reaction has a well-defined pH optimum at 

 neutrality. This is shown in Fig. 6. A study of the temperature-dependence 



10 20 30 40 50 

 Incubation time (min) 



Fig. 5. The specific effect of sodium ions in promoting the transport of free 

 [i--'*C]-alanine into the amino acid "pool" of the isolated cell nucleus. The 

 specific activity of the acid-soluble nuclear extracts (see text) is plotted against 

 the time of incubation at 37°. 



of the reaction reveals the optimum in the physiological temperature range 

 (38°-40°) (Fig. 7). A comparison of the rates of alanine transport at 10° 

 and 20° (0-83 and i -6 m^moles of L-alanine per 10 min. per 20 mg. nuclei) 

 shows that the velocity of the reaction has doubled for a 10° rise in tem- 

 perature. This temperature coefficient (Ojo = 2) indicates that amino acid 

 transport into the nucleus involves a chemical reaction and is not simply a 

 result of diffusion. 



It has already been pointed out that even after short periods of incuba- 

 tion, the [^^C]-alanine concentration within the nuclei may exceed that 

 of the incubation medium. This transport and accumulation of an amino 

 acid against a concentration gradient would be expected to require 

 the expenditure of energy. In this connection there is some evidence 



