Microtubular protein in its polymerized or nonpolymerized states differentially modulates in vitro and intracellular fluxes catalyzed by enzymes of carbon metabolism

S. Cortassa, M. A. Aon, A. Cáceres

Research output: Contribution to journalArticlepeer-review

Abstract

The fluxes through HK/G6PDH and PK/LDH coupled‐enzymatic reactions were quantified in the presence of physiological concentrations (1–15 μM) of polymerized or non‐polymerized microtubular protein (MTP) from rat brain and in a permeabilized yeast cell system. In vitro enzymatic fluxes were increased by either polymerized or nonpolymerized brain MTP mainly in the lower range of MTP concentration. At fixed MTP concentrations in the flux stimulatory range of HK/G6PDH (1 mg/ml MTP) or PK/LDH (0.4 mg/ml MTP), a hyperbolic and sigmoidal response to NADP and PEP, respectively, was detected. That dependence varied according to the polymeric status of MTP. The specificity of the phenomenon observed in vitro, was tested for the PK/LDH and HK/G6PDH enzymatic couples in the presence of neutral polymers such as glycogen (⩽ 10 mg/ml), poly(ethylene glycol) (up to 10% w/w) or G‐actin (⩽ 1 mg/ml). In permeabilized Saccharomyces cerevisiae cells, the PK‐catalyzed flux was sensitive to microtubule disruption by nocodazole (15 μg/ml). The HK/G6PDH system was not affected by nocodazole showing values of kinetic parameters close to those obtained in vitro in the presence of polymerized brain MTP. Indirect immunofluorescence with specific antibodies against tubulin allowed to confirm the microtubules disruption in the presence of nocodazole in permeabilized yeast cells under the same conditions in which enzymes were assayed intracellularly. The experimental evidence is in agreement with the observed phenomenon of increase in fluxes in the enzymatic reactions assayed to be specifically induced by MTP either in vitro or in situ. The results presented are discussed in terms of the assembly of large supramolecular structures as a supraregulatory mechanism of synchronization of systemic cellular processes such as metabolic fluxes. © 1994 Wiley‐Liss, Inc.

Original languageEnglish (US)
Pages (from-to)120-132
Number of pages13
JournalJournal of cellular biochemistry
Volume55
Issue number1
DOIs
StatePublished - May 1994

Keywords

  • carbon metabolism
  • flux regulation
  • microtubular protein
  • microtubules
  • permeabilized yeast cells

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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