VOL. 12 (1953) AMINO ACID INTERACTIONS IN STRICT ANAEROBES 



115 



Effects of Pyruvate, a-ketoglutarate and thiol compounds on resting CI. sporogenes after 

 exposure to air or hydrogen peroxide 



Exposure of resting CI. sporogenes to air at 37° causes a fall in its ability to oxidise 

 L-alanine (see Table XII) and the addition of pyruvate has but little stimulant effect. 

 If, however, the exposure of CI. sporogenes to air takes place in the presence of o.oi M 

 pyruvate or a-ketoglutarate, its subsequent ability to oxidase alanine is much increased 

 and the stimulating action of the presence of pyruvate is again noticeable (Table XII). 



TABLE XII 



EFFECTS OF PYRUVATE AND a-KETOGLUTARATE ON ACTIVITY OF RESTING CI. SpOVOgeUeS 



Warburg manometer vessels contained i ml suspension of CI. sporogenes (in 0.15 M KCl) in 

 0.02 M phosphate buffer pH 7.3. Total vol. 3.2 ml. L-Alanine = 0.02 M. Pyruvate = o.oi M. 



Contents of vessel 



Conditions of exposure of 

 CI. sporogenes 



flM oxygen 

 absorbed 



liberated 



Alanine 

 Alanine 



Alanine + Pyruvate 



Alanine 



Alanine + Pyruvate 



Alanine 



Alanine + Pyruvate 



Freshly prepared organism used immediately. 10.7 



I ml organism diluted with 2 ml water, exposed 



to air at 37° for 40', centrifuged and resus- 



pended in i ml 0.15 M KCl 4.3 



I ml organism diluted with 2 ml water, exposed 



to air at 37° for 40', centrifuged and resus- 



pended in i ml 0.15 M KCl 5.7 



I ml organism diluted with 2 ml sodium 



pyruvate soln. (o.oi M) exposed to air at 37° 



for 40 min centrifuged and resuspended in j 



ml 0.15 M KCl 8.9 



I ml organism diluted with 2 ml sodium 



pyruvate soln. (o.oi M) exposed to air at 37° 



for 40' centrifuged and resuspended in i ml 



0.15M KCl 16.6 



I ml organism diluted with 2 ml sodium 



a-ketoglutarate soln. (o.oi M) exposed to air 



at 37 ° for 1 o', centrifuged and resuspended i ml 



0.15 M KCl 7.7 



I ml organism diluted with 2 ml sodium 



aketoglutarate soln. (o.oi M) exposed to air 



at 37° for 10', centrifuged and resuspended in i 



ml 0.15 M KCl 14.0 



15.0 

 6.7 



7-7 



11.8 

 20.5 

 11.6 

 13.8 



This phenomenon may be explained as being due to a toxic effect of oxygen on the 

 amino acid oxidase system, that is obtained even in the absence of the amino acid, when 

 there is no perceptible absorption of oxygen. The protective effects of pyruvate and 

 a-ketoglutarate may be due either to destruction of traces of hydrogen peroxide or to a 

 reversible linkage with thiol groups important in the oxidation of alanine, thus preserving 

 them from irreversible oxidation by oxygen (or hydrogen peroxide). The fact that thiol 

 groups are involved in the aerobic oxidation of L-alanine by CI. sporogenes is indicated 

 by the inhibitory effect of phenyl arsenoxide thioglycoUate (200 /xg/vessel) which 

 diminishes the rate of alanine oxidation by 81 %. 



The considerable diminution of the rate of anaerobic amino acid (alanine-glycine, 

 interaction brought about by exposure of resting CI. sporogenes to air at 0° (Tables XI) 

 XII) may be prevented by the presence of sodium thioglycoUate. Typical results are 

 shown in Table XIII. This result would be expected it thioglycoUate prevents access of 

 oxygen to the organism by its preferential oxidation. 



References p. 120. 



