STRUCTURE‐FUNCTION RELATIONSHIPS IN PYRUVATE DECARBOXYLASE OF YEAST AND WHEAT GERM*<sup>†</sup>

Ullrich, Johannes

doi:10.1111/j.1749-6632.1982.tb31203.x

Cited by 42 publications

(30 citation statements)

References 68 publications

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“…(14) and for PDC of yeast (19). The observation of two subunits of slightly differing mol wt for both maize PDCs was also consistent with the values reported for yeast PDC (23). This finding differed from the single mol wt species reported for PDC from sweet potato (14).…”

Section: Methodscontrasting

confidence: 52%

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Pyruvate Decarboxylase from Zea mays L.

Lee

Langston-Unkefer²

1985

Plant Physiol.

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Pyruvate decarboxylase (PDC) was purified from mature, dry maize kernels and from roots of anaerobically treated maize seedlings and partially characterzed. PDC was purified to a specific activity of 96 units per milligram protein from kernels and to 41 units per milligram protein from root. The subunit molecular masses were estimated to be 61,000 and 60,000 for kernel PDC and 59,000 and 58,000 for root PDC. The pH optimum for each enzyme was 5.8. Since the pH optimum is nearly one pH unit below the value reported for the cytoplasm of anaerobically metabolizing maize roots (pH 6.7 1 0.2), we investigated the effects of pH 5.8 and 6.6 on the cooperative kinetics observed for PDC from each source. The maximum Hill coefficients (nH) were much greater at each pH for the kernel PDC (pH 5.8, nH = 2.5 and pH 6.6, nH = 3.2) than for the root PDC (pH 5.8, nH = 14A and pH 6.6, nH = 1.8). The cooperative kinetics observed with respect to pyruvate were asymmetric. Potassium inhibited maize PDC and was competitive with pyruvate (root PDC K,= 16 millimolar and kernel PDC KJ= 10 millimolar).Pyruvate decarboxylase, PDC3 (EC 4.1.1.1) catalyzes the decarboxylation of pyruvate to acetaldehyde; ADH (EC 1.1.1.1) then catalyzes the reduction ofthe acetaldehydeto ethanol. These two enzymes thus catalyze a pathway in which NAD+ is regenerated under anaerobic conditions. Pyruvate, the first substrate of this pathway, occurs at a major branch point in glycolysis between anaerobic and aerobic metabolism, and the regulation between these two metabolic routes is likely to occur both at the level of PDC synthesis and of PDC activity. The syntheses of PDC and ADH are specifically enhanced under anaerobic conditions (9, 18), and significant increases in the activities of these enzymes are observed (9, 18). One functional mechanism proposed to control this anaerobic pathway is the regulating effect of cytoplasmic pH on PDC activity (7). PDC is inactive at the pH ofaerobically metabolizing tissues. However, the cytoplasmic '

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Section: Methodscontrasting

confidence: 52%

“…The lag phase observed with maize PDC is also characteristic of PDC from yeast (23). We have investigated the lag phase, and the results of this investigation will be reported elsewhere.…”

Section: Methodsmentioning

confidence: 99%

Pyruvate Decarboxylase from Zea mays L.

Lee

Langston-Unkefer²

1985

Plant Physiol.

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“…Despite numerous attempts to probe the active sites of TPP-dependent enzymes by means of chemical modification [22][23][24][25][26], no amino acid sequences have yet been identified, although it is widely accepted that the TPP-binding sites must be hydrophobic in character [27][28][29][30]. Our identification of a common sequence motif in TPPdependent enzymes catalysing a wide variety of chemical reactions now opens the way to a systematic study of this motif by means of sitedirected mutagenesis and protein engineering.…”

Section: Discussionmentioning

confidence: 99%

A common structural motif in thiamin pyrophosphate‐binding enzymes

1989

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The amino acid sequences of a wide range of enzymes that utilize thiamin pyrophosphate (TPP) as cofactor have been compared. A common sequence motif approximately 30 residues in length was detected, beginning with the highly conserved sequence -GDG-and concluding with the highly conserved sequence -NN-. Secondary structure predictions suggest that the motif may adopt a flail fold. The same motif was recognised in the primary structure of a protein deduced from the DNA sequence of a hitherto unassigned open reading frame of Rhodobacter capsulata. This putative protein exhibits additional homology with some but not all of the TPP-binding enzymes.

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“…The enzyme of Saccharomyces cerevisiae is composed of four identical subunits of molecular mass about 62 kDa each [2, 31. The synthesis of pyruvate decarboxylase is induced by high glucose concentration at the transcriptional level [4 -61.…”

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confidence: 99%

Autoregulation may control the expression of yeast pyruvate decarboxylase structural genes PDC1 and PDC5

Hohmann

Cederberg

1990

European Journal of Biochemistry

152

124

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Recently we deleted the pyruvate decarboxylase structural gene PDCl from the genome of the yeast Saccharomyces cerevisiae. The pdcl deletion mutants had pyruvate decarboxylase activity due to the presence of a second structural gene [Schaaff, I., Green, J. B. A,, Gozalbo, D. & Hohmann, S. (1989) Curr. Genet. 15, [75][76][77][78][79][80][81]. We cloned and sequenced this gene which we call PDCS. The predicted amino acid sequences of PDCl and PDCS are 88% identical.Deletion of PDCS did not cause any decrease in the specific pyruvate decarboxylase activity whilepdcl deletion mutants had 80% of the wild-type activity. Deletion mutants lacking both PDCl and PDCS did not show any detectable pyruvate decarboxylase activity in vitro and were unable to ferment glucose. This indicates that PDCl and PDCS are the only structural genes for pyruvate decarboxylase in yeast.The PDCS isoenzyme showed a slightly higher K , value for its substrate pyruvate than the PDCI product (PDCS: K, = 8 mM; PDCI : K, = 5 mM), as measured in crude extract of pdcl and pdc5 deletion mutants, respectively.PDCS is only expressed in pdcl deletion mutants. No mRNA transcribed from PDCS could be detected in wild-type cells. Thus, in addition to the control by glucose induction, pyruvate decarboxylase activity seems to be subject to autoregulation. Similar phenomena have been described previously for tubulin, histones and a ribosomal protein but not for metabolic enzymes.Pyruvate decarboxylase is the key enzyme of alcoholic fermentation since it cleaves pyruvate, the end product of glycolysis, to CO, and acetaldehyde [1]. .2).Note. The nucleotide sequence of the PDCS DNA sequence published here has been deposited with the EMBL/GenBank/DDBJ Nucleotide Sequence data bank and will be availablc under the accession number XI 5668. Surprisingly, a pdcl deletion mutation conferred a different phenotype than thepdcl-8 mutation described by Schmitt and Zimmermann [7]. The deletion mutants fermented glucose but they had about 50% of the wild-type pyruvate decarboxylase activity [12]. This activity was most probably due to a second gene encoding pyruvate decarboxylase which is very similar to PDCI since the pdcl deletion mutants still had a full-length mRNA hybridizing to PDCl probes in Northern blot analysis [12].Here we report the cloning and molecular analysis of PDC5, the second pyruvate decarboxylase structural gene. This gene seems to be not, or only poorly, expressed in wildtype cells but is activated in the pdcl deletion mutants. MATERIALS AND METHODS Yeast strainsStrain 1.1.-6B

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STRUCTURE‐FUNCTION RELATIONSHIPS IN PYRUVATE DECARBOXYLASE OF YEAST AND WHEAT GERM*^†

Cited by 42 publications

References 68 publications

Pyruvate Decarboxylase from Zea mays L.

Pyruvate Decarboxylase from Zea mays L.

A common structural motif in thiamin pyrophosphate‐binding enzymes

Autoregulation may control the expression of yeast pyruvate decarboxylase structural genes PDC1 and PDC5

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STRUCTURE‐FUNCTION RELATIONSHIPS IN PYRUVATE DECARBOXYLASE OF YEAST AND WHEAT GERM*†

Cited by 42 publications

References 68 publications

Pyruvate Decarboxylase from Zea mays L.

Pyruvate Decarboxylase from Zea mays L.

A common structural motif in thiamin pyrophosphate‐binding enzymes

Autoregulation may control the expression of yeast pyruvate decarboxylase structural genes PDC1 and PDC5

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STRUCTURE‐FUNCTION RELATIONSHIPS IN PYRUVATE DECARBOXYLASE OF YEAST AND WHEAT GERM*^†