Biochemical and genetic characterization of pyruvate decarboxylase from the yeast Candida zemplinina

CONTI, FRANCISCO; MARÍA LAURA RAYMOND EDER; ALBERTO LUIS ROSA

Paraná

Congreso; Reunión Anual Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2018

Pyruvate decarboxylase (PDC) is the enzyme responsible for the non-oxidative decarboxylation of pyruvate to acetaldehyde and carbon dioxide. PDC plays an essential role in the yeast fermentation pathway leading to ethanol production. Saccharomyces cerevisiae has three active PDC isoenzymes (PDC1, PDC5 and PDC6) which, similar to PDC from other yeasts species, exhibit substrate activation. Here we present the first biochemical and genetic characterization of PDC from the yeast C. zemplinina (syn., Starmerella bacillaris). CzPDC activity was readily detected in total cell lysates from C. zemplinina type strain 9494 as well as from several indigenous C. zemplinina isolates from spontaneously fermenting grape musts. Characterization of kinetic parameters of PDC1 and CzPDC, through steady state measurement, showed that CzPDC does not present substrate activation but a standard Michaelis-Menten kinetic behavior. In silico analyses of the recently available complete genome sequence of several S. bacillaris isolates, including C. zemplinina type strain 9494, suggest the existence of a single gene encoding CzPDC. Further analyses of the conceptually translated amino acid sequence of CzPDC revealed that critical amino acids at the PDC catalytic site are highly conserved among CzPDC and S. cerevisiae PDC1, PDC5 and PDC6. However, critical residues C221, H92 and H225, involved in PDC substrate activation and present in PDCs from several yeast species, are absent in the C. zemplinina enzyme. This observation supports the results obtained from the kinetic analyses of CzPDC. This report represents the first description of a yeast PDC lacking substrate activation.