SIDNEY SHJIRIN AND NATHAN O. KAPLAN 145 



TABLE 1 



ProIM'RI ll.S ()!■ fS AND iv DPN 



Properties 0-DPN a-DPN 



Enzymatic activity active inactive 



Ciiemical reduction Xmax 340 iriyu X„,ax 345 m/u 



Cyanide addition X,„„x 325 m^ Xmax 332 m/x 



Optical rotation — 34 . 8 -1-14.3 



Optical rotation of corresponding — 38.3 -)- 58.2 



nicotinaniidc mononucleotides 



ni/x upon excitation 345 ni/t. However, when the reduced a isomer 

 WAS excited at 260 m^, a wavelength at which 95 per cent of the 

 light is absorbed by the adenine, the fluorescence intensity at 465 m^ 

 is only a fraction of that obtained from direct excitation of the 

 dihydronicotinamide (Fig. 1) (20). These results are in contrast to 

 those obtained with the beta, or enzymatically active, isomer of DPNH, 

 in ^\•hich 260 ni/^ excitation results in the fluorescence of the di- 

 hydronicotinamide at 462 mfx with an intensity equal to that ob- 

 tained from 340 m^ excitation. The decreased efficiency of intra- 

 molecular energy transfer suggests that the two components of the 

 a-dinucleotide are not maintained in optimum conformations to allow 

 for interaction. 



Another variation in the structure of the DPNH molecule which 

 may aid in elucidating the nature of the forces facilitating intra- 

 molecular interactions is the use of a thioamide group in place of 

 the carboxamide of nicotinamide. Unlike DPN, which is transparent 

 above 260-270 m^x, the corresponding thionicotinamide analogue has 

 a pronounced absorption maximum at 300 m^, as shown by curve A 

 in Fig. 2. The position of this maxinumi conforms to the suggestion 

 made by Braude (5) that replacement of an oxygen by a sulfur 

 auxochromic group in many chromophores results in a shift of the 

 pertinent absorption maximum by 40-50 m^ toward longer wave- 

 lengths, with an increase in the extinction coefficient. The generaliza- 

 tion is further borne out by the spectrum of the enzymatically re- 

 duced analogue, shown in curve B. The absorption band has its 

 maximum at 398 m^ (l()-3 g — 8.9) and the prominent band at 

 300 m/x has become an inflection point with its maxinunn at 295 

 mix. The specific absorption band of the thiocarbonyl group permits 

 an examination of the participation of this functional group in intra- 

 molecular interactions. 



We postulate the structine of thionicotinamide-DPNH to be that 

 shown in Fig. 3, in ^vhich the 6-amino group of adenine is hydrogen- 



