236 BIOCHEMISTRY OF BACTERIAL LUMINESCENCE 



type enzyme and the flavin then is oxidized in the luminescent flavin 

 oxidase. According to this picture aldehyde functions in this sequence 

 by forming an oxygen adduct ( Jockusch, 1949; McDowell and Thomas, 

 1949; Wittig and Pieper, 1941), which oxidizes the flavin forming in 

 essence a peroxide aldehyde addition product.* This peroxy aldehyde 

 then oxidizes another flavin or, as mentioned alternatively, peroxide 

 with the subsequent emission of light. 



The slowest half-rise times are elicted by the DPN-substrate 

 reactions; the intermediate rise times occur when the DPNH2-FMN 

 reaction is the starting point; and the more rapid rise times are 

 characteristic of the main luminescent reactions involving a somewhat 

 complicate mechanism of FMNH2 oxidation. 



The pressure studies give some clue as to the site of action of the 

 aldehyde. Among the observations which must» be fit into any 

 scheme which purports to present a unified picture are included the 

 fact that reduced flavin requires aldehyde for maximal luminous 

 oxidation, that the pressure effects are not observable in the absence 

 of aldehyde, and that limiting amounts of aldehyde produce an 

 anomalous type of pressure response. Since reduced flavin is easily 

 auto-oxidizable by oxygen the lack of effect of KCF on the rate of 

 respiration at high oxygen tensions is simply due to the fact that 

 auto-oxidation of the flavins maintains the respiratory rate. 



When aldehyde is added, however, the pathway is considerably 

 diflFerent. The flavin is oxidized by the aldehyde oxygen addition 

 product, and the pool size of reduced FMN is diminished. The inter- 

 mediate "D" represents the hypothetical aldehyde peroxide addition 

 product, which is formed only in the presence of KCF and reduced 

 flavin. If the concentration of aldehyde is nil or vanishingly small the 

 amount of D which accumulates is minescule and, therefore, since 

 the reaction B to D is limiting, the rate of light output cannot be 

 accelerated by pressure. Moreover, since the pool size of FMNHo will 

 remain large, any B to D enzyme remaining will be essentially 



* We have recently synthesized in our laboratory the crystalline peroxide- 

 nonaldehyde addition product (alpha-oxynonylhydroperoxide) and find its 

 luminescence potentiating effect to be several times that of the free aldehyde 

 on a molar basis. Although this observation does not necessarily support the 

 scheme as set forth, it is interesting that this derivative is more effective in 

 promoting luminescence than the free aldehyde. 



