Metabolic changes in Saccharomyces cerevisiae strains lacking citrate synthases

Area of Science:

• Biochemistry
• Yeast Metabolism
• Enzymology

Background:

• Saccharomyces cerevisiae possesses two citrate synthase isoenzymes: mitochondrial (CS1) and cytosolic (CS2).
• Understanding the specific roles and metabolic consequences of these isoenzymes is crucial for comprehending yeast energy production and biosynthesis.

Purpose of the Study:

• To investigate the metabolic effects of deleting CS1, CS2, or both isoenzymes in Saccharomyces cerevisiae.
• To elucidate the impact of citrate synthase deficiency on yeast growth, substrate utilization, and related enzyme activities.

Main Methods:

• Construction and characterization of CS1-, CS2-, and CS1-CS2- mutant strains of Saccharomyces cerevisiae.
• Growth rate analysis on various carbon sources (galactose, glycerol, lactate, pyruvate, glutamate, acetate).
• Measurement of intracellular metabolite concentrations (citrate, malate) and 14CO2 production from labeled substrates.
• Enzymatic assays for citric acid cycle and electron transport chain components.

Main Results:

• CS1- mutants exhibited prolonged lag phases on non-fermentable carbon sources and failed to grow on acetate.
• Absence of CS1 led to increased intracellular citrate and malate levels when grown on lactate or pyruvate.
• Reduced 14CO2 production from acetate and glutamate in CS1- and CS1-CS2- mutants indicated impaired carbon flux.
• Decreased activities of pyruvate dehydrogenase complex, alpha-ketoglutarate dehydrogenase complex, and aconitase were observed in CS1- mutants.

Conclusions:

• The absence of mitochondrial citrate synthase (CS1) has significant metabolic consequences beyond its direct enzymatic role.
• CS1 deficiency impacts yeast growth, carbon metabolism, and the activity of other key enzymes in central metabolic pathways.
• These findings highlight the complex interplay of citrate synthase isoenzymes in yeast metabolic regulation.