An improved procedure was developed for the isolation of pyruvate decarboxylase from wheat germ. Its final step, an electrophoresis of the native apoenzyme in concave pore gradient polyacrylamide gels, followed by superficial activity-staining, produced two bands of different molecular masses and chain compositions. The high-molecular-mass band occurred in low quantity and consisted of, probably eight, apparently identical chains of Mr = 33,000, as judged from sodium dodecyl sulfate electrophoreses. The low-molecular-mass band contained two types of chains with Mr alpha = 63,000-65,000 and Mr beta = 61,000-62,000. The N termini of both chains were threonine, whereas their C-terminal sequences were different: alpha, -(Val)-(Ser)-(Ala)-Leu; beta, -(His)-(Asp)-(Ala)-Ser. Their amino acid composition was too different to be compatible with our original concept of one chain being produced from the other by proteolytic shortening. Limited proteolysis by Staphylococcus aureus V8 proteinase yielded peptides partly identical size and partly quite different. In all properties investigated, the low-molecular-mass enzyme largely resembled yeast pyruvate decarboxylase; the holoenzyme appeared to possess (alpha beta)2 structure, the apoenzyme alpha beta. SH reagents inactivated the enzyme. Binding and fluorescence of 2-p-toluidinonaphthalene-6-sulfonate indicated a similar lipophilicity of the active site as found earlier for the yeast enzyme. 2-Hydroxy-5-nitrobenzyl modification of exposed tryptophan residues left the holoenzyme intact, but in the apoenzyme it destroyed most of the cofactor-binding ability and hence the activity. The strength of cofactor binding and the maximal specific activity were found somewhat lower than in yeast pyruvate decarboxylase.

Amino Acids↗

PubMed1983

Activity stain for pyruvate decarboxylase in polyacrylamide gels.

A method for the localization of pyruvate decarboxylase bands in polyacrylamide gels after electrophoresis using 1,2-dianilinoethane in dilute acetic acid as reagent for acetaldehyde formed from pyruvate by the holoenzyme is described. A white condensation product forms in the bands and precipitates within a few minutes. The more or less opaque bands can be viewed and photographed against a dark background and scanned in a densitometer. The detection limit is at about 10 mU pyruvate decarboxylase when the gels are bathed in the staining solution for 20 min. Several other methods were tested and failed to produce satisfactory results.

Carboxy-Lyases↗

PubMed1982

Influence of prolactin on metabolism and energy production in perfused corpus luteum bearing bovine ovaries.

Eight bovine ovaries with a corpus luteum were perfused for 4 h in a haemoglobin-free semi-synthetic perfusion medium in a closed circuit. After an initial prolactin (PRL)-free perfusion phase, 4.7 ng/ml ovine PRL was added in the 1st h, followed by 47 ng/ml in the 2nd h and 470 ng/ml in the 3rd h. Glucose and oxygen consumption and the production of lactate, pyruvate and CO2 were measured, while perfusion pressure and pH-value were recorded continuously. Under the influence of PRL anaerobic glucose metabolism was stimulated by 40.5% and oxidative phosphorylation was inhibited. Energy production from aerobic glucose metabolism rose by only 0.25%. Unlike PRL, Human Menopausal Gonadotropin (hMG) and Human Chorionic Gonadotropin (hCG) stimulated aerobic metabolism. This may indicate that PRL is the "older" hormone in phylleogenetic terms.

Animals↗

PubMed1982

Structure-function relationships in pyruvate decarboxylase of yeast and wheat germ.

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Acetylation↗

PubMed1976

[Clinical symptoms of acoustic neurinoma. A contribution to the diagnosis with reference to neurologic and otologic findings].

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Adult↗

PubMed1975

[Stellate ganglion block under EEG control in the treatment of complicated cervical migraine (author's transl)].

Complicated migraine is regarded as a cerebrovascular syndrome of functional character. In the case of a young man with cervical migraine associated with paroxysmal neurological disorders and persisting EEG abnormalities, stellate ganglion blocks were performed under EEG control. A transient amelioration of the pathological EEG pattern was observed after each stellate ganglion block and after several treatments the EEG was normal and the migraine attacks disappeared. The pathophysiology of the effect of stellate ganglion block on cerebral blood flow is discussed.

Adult↗

PubMed1975