Monfoort, C.H.

Utrecht University Repository

572 BIOCHIMICA ET BIOPHYSICA ACTA VOL. 9 (1952) Short Communications and Preliminary Notes ON THE RAT IO OF  THIAMINE PYROPHOSPHATE CONTENT AND RATE OF  SUCCIN IC  SEMIALDEHYDE PRODUCTION IN  HOMOGENATES OF  VARIOUS MUSCLES OF  NORMAL AND THIAMINE-DEF IC IENT P IGEONS by C. H. MONFOORT Laboratory [or Physiological Chemistry, The University, Utrecht (Netherlands) In  a previous communication 1 we were able to show that large differences exist between various types of muscles regarding (I) the rate of production of acetoin from pyruvate added to homogenates, and (2) the rate at which the acetoin-producing enzyme, the pyruvic decarboxylase, disappears from these muscles upon thiamine privation. The experiments were carried out with homogenates of the large breast muscle (pectoralis major), the muscle of the left heart ventricle and the leg muscles of normal pigeons and of pigeons which had been on a thiamine-free diet for 4 and 12 days. For details see our earlier communication s . In this preliminary note the results will be given of the investigation along similar lines of another thiamine pyrophosphate (TPP) dependent enzyme, the a-ketoglutaric decarboxylase, which catalyses the reaction a-ketoglutaric acid -> succinic semialdehyde + CO s. The succinic semialdehyde (SSA) formed from a-ketoglutarate added to the muscle homogenate was determined according to a method based on the principle given by OCHOA s. The results are assembled in Table I. Q = the amount of SSA formed per 20o mg tissue per 3 hours, divided by the amount of TPP  per 2oo mg tissue. The following conclusions may be drawn (QB, QH and QL denote Q for breast muscle, heart muscle and leg muscle, respectively): I. Normal pigeons. The rate of SSA formation declines in the same order as the rate of acetoin formation (see x) and as the TPP  contents: breast muscle > heart muscle > leg muscle. QB = QH > QL. This means- -prov ided that the assumptions, mentioned in Ref. i are va l id - -  that  in breast muscle and heart muscle the same percentage of the total TPP  is present as the pros- thetic group of a-ketoglutaric decarboxylase, and that this percentage is higher than that in leg muscle. 2. Influence o/ thiamine deficiency. The TPP  content decreased uring the whole period of thiamine privation in every type of muscle to the same extent as in our previous experiments 1. In the case of the breast muscle the production of SSA also decreases during the first 4 days of deficiency, but it remains constant during the next 8 days. The ratios of the changes of TPP content and SSA production are such, that  ~B remains constant during the first 4 days and then increases very considerably. In heart muscle, however, there is no change of a-ketoglutaric decarboxylase activity in the first 4 days, while it diminishes lightly in the next 8 days. This implies that ~H is about doubled in both periods. In leg muscle there is hardly any influence of thiamine deficiency on the activity of the a-keto- glutaric decarboxylase. Therefore ~L increases considerably. In general one may conclude that  with the exception of breast muscle during the first 4 days of deficiency, where the pyruvic decarboxylase appears to be little affected by the disappearance of TPP  from the tissue, the a-ketoglutaric decarboxylase is much better maintained uring deficiency than the pyruvic decarboxylase. VOL. 9 (1952) SHORT COMMUNICATIONS, PRELIMINARY NOTES 573 TABLE I TItlAMINE PYROPHOSPHATE CONTENT AND SUCCINIC SEMIALDEFIYDE PRODUCTION In HOMOGENATES OF PIGEON MUSCLE B = breast muscle, H = heart muscle, L = leg muscle. TPP  expressed in y per 2oo mg, SSA in 7 per 200 mg in 3 hrs. <) = 7 SSA per 20o mg in 3 hrs y TPP  per 200 nlg Numbers of muscles examined: Normal 4 d. def. a a d. def. B 7 9 9 H 7 8 9 L 7 8 9 All standard eviations mentioned are standard eviations of the means. Normal 4 days def. i2 days def. TPP  SSA Q TPP SSA Q TPP " SSA Q B I '5°~° ' I °  728£47 486±13 0.94±0.04 473±37 5°3 q-2I 0.62+0.06 493±4 ° 800±62 H 1.29-[-o.o 7 588~-37 462±27 0.77±o.05 62oJ=49 825J-82 0.304-0.07 466±17 1637±125 L o.45~o.o 4 I I7~I6  255±21 o.22-co.oi 78±3 3554-16 o.14+o.o2 85~:IO 628+68 P values calculated according to "STUDENT" 's method: B PQ(N-4d.) > o.i PQ(i2d.-N) < o.ooi PQ(~2d.-4d.) < o.ooi PSSA(N-4d.) < o.ooi PSSA(4d.-I2d,) ~ O.I H mQ(4d.-N) < O.OOI PQ(I2d,-N) < o.ooi PQ(I2d.-4d.) < 0.00I PSSA(N_4d.) > o.I PSSA(N-I2d.) ~ o.oi PSSA(4d.-x2d.) ~ o.oi L Po(4d.-N) < 0.0i PQ(i2d.-N) < 0.00i PQ(i2d.-4d.) ~ 0.0i PSSA(N 4d.) < 0-05 PSSA(N-I~d.) ~ 0.05 PSSA(I2d.-4d.) ~ 0.05 Increase and decrease of Q indicate that  the enzyme studied disappears more slowly or more rapidly, respectively, than the other TPP  dependent enzymes collectively. If Q is constant he enzyme studied and the latter enzymes disappear at equal rates. Upon comparison of the change of the ~ values for a-ketoglutarie acid decarboxylation with the change of Q values for pyruvic acid decarboxylation (seel), it appears that  there exist great differences between the disappearance of a-ketoglutaric decarboxylase and pyruvic decarboxylase in relation to the loss of TPP. During the first 4 days of deficiency the pyruvic decarboxylase disappears more slowly than the other TPP-dependent enzymes collectively (including the a-ketoglutaric decarboxylase). The same applies to the disappearance of a-ketoglutaric decarboxylase and the other TPP-dependent enzymes collectively (including the pyruvic decarboxylase). During the next 8 days of deficiency, however, the pyruvic decarboxylase disappears at a higher rate than the other TPP-dependent enzymes collectively, while the a-ketoglutaric decarboxylase continues to disappear at a lower, occasionally at a much lower, rate. This work forms part  of the investigations on the metabolism and physiological function of thiamine carried out by H. G. K. WESTENBRINK and collaborators. REFERENCES 1 C. H. MONFOORT, Bioch im.  B iophys .  Acta, 9 (1952) 33 I. 2 C. H. MONFOORT, Bioch im.  B iophys .  Acta, 8 (I952) 589. 3 S. OCHOA, J .  Biol .  Chem., 155 (1944) 87. Received September 9th, 1952 

On the ratio of thiamine pyrophosphate content and rate of succinic semialdehyde production in homogenates of various muscles of normal and thiamine-deficient pigeons

On the ratio of thiamine pyrophosphate content and rate of succinic semialdehyde production in homogenates of various muscles of normal and thiamine-deficient pigeons

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