112 



R. MAMELAK, J. H. QUASTEL 



VOL. 12 (1953) 



TABLE IX 



ACETYLATION OF SULFANILAMIDE DURING AMINO ACID INTERACTIONS IN PRESENCE OF 



CI. sporogenes extract 



Warburg manometer vessels contained i ml extract of CI. sporogenes (in i % thioglycollate and 

 o.oi M phosphate) and 0.5 ml extract of pigeon liver (500 mg acetone pigeon liver powder suspended 

 in 5 ml 0.2 M KF and supernatant used), 0.028 M NaHC03 present. Gas = 93% Ng + 7% COg. 

 Temp.: 37°. Time 75'. DPN = 3 mg/vessel. Sulfanilamide = 200 ^<g/vessel (Expts. 1,3) and 400 

 ^<g/vessel (Expt. 2). Amino Acids = 0.02 M (L-form). 



Expt. 



Contents of vessel 



JilM CO, 



[Ig sulfanilamide 

 acetyl ated 



Bact. extr. + Pigeon liver extr. 



Bact. extr. + Alanine + Proline 



Pigeon liver Extr. + Alanine + Proline 



Bact. extr. + Pigeon liver Extr. + Alanine + Proline 



Bact. ext. + Pigeon liver extract 



Bact. extr. + Pigeon liver extr. + Alanine + Proline 



Bact. extr. + Pigeon liver extr. + 0.005 M Arsenite 



Bact. extr. + Pigeon liver extr. + Alanine + Proline + 



0.005 ^^ Arsenite 

 Pigeon liver extr. + Pyruvate (.01 M) + ProUne 

 Bact. extr. + Pigeon liver extr. + Pyruvate (o.oi M) + 



Proline 

 Bact. extr. + Pigeon liver Extr. 



Alanine + DPN + H,0 = Pyruvate + NH3 + DPNHg 

 Pyruvate + CoA + DPN ^ Acetyl-CoA + CO^ + DPNH2 

 Acetyl-CoA -f Sulfanilamide — Acetylsulfanilamide + CoA 



Such a conclusion is supported by the experimental evidence that CoA is present in 



extracts of CI. sporogenes (Nisman and Mager^^)^ 



Effects of pyruvate on alanine oxidation by resting CI. sporogenes 



L- Alanine is oxidised aerobically by resting CI. sporogenes, a vigorous rate of oxygen 

 uptake, which falls off markedly with time, taking place. If pyruvate is an intermediate, 

 as would be anticipated from the evidence given above, it would be expected that either 

 pyruvate would be oxidised aerobically as vigorously as alanine or that it would accumu- 

 late during the oxidation. In fact, however, the rate of oxygen uptake by resting CI. 

 sporogenes in presence of pyruvate is remarkably feeble. Typical results for alanine and 

 pyruvate are given in Table X. Moreover, estimation of the pyruvic acid indicates little 

 or no disappearance in presence of CI. sporogenes under aerobic conditions. During aerobic 

 oxidation of alanine there is no accumulation of pyruvate, a fact noted by all previous 

 workers. The problem arose, therefore, as to why pyruvate does not accumulate, if it 

 does not undergo aerobic oxidation by CI. sporogenes. 



The following observations throw light on this phenomenon. The addition of pyruvate 

 to DL-alanine markedly extends the interval during which vigorous aerobic oxidation of 

 alanine takes place, so that in effect the oxygen uptake by CI. sporogenes due to the alani- 

 ne after a lengthy interval is greatly increased by the presence of pyruvate. Typical results 

 are given in Table X. This phenomenon is also observed when a-ketoglutarate is sub- 

 stituted for pyruvate. During the period of marked stimulation of oxygen uptake due to 

 the ketonic acid, little or none of the latter disappears. There is, however, a large increase 

 of the rate of disappearance of alanine and of the rate of liberation of ammonia (see 



References p. 120. 



