[CaCO3 stimulates alpha-ketoglutarate accumulation during pyruvate fermentation by Torulopsis glabrata].
A large amount of alpha-ketoglutarate (alpha-KG) (6.8 g/L) was accumulated in flask culture when CaCO3 was used as a buffering agent in the production of pyruvate by multi-vitamin auxotrophic yeast Torulopsis glabrata CCTCC M202019. In a 5 L jar-fermentor, less alpha-KG (1.3 g/L) was produced when NaOH was used to adjust the pH, while more alpha-KG (11.5 g/L) detected when CaCO3 was used as the buffer. In the latter case, the molar carbon ratio of pyruvate to alpha-KG (C(PYR)/ CalphaKG) was similar to that obtained in flask culture, suggesting the accumulation of alpha-ketoglutarate was related to the addition of CaCO3. Furthermore, it was found that: (1) delaying the addition time of CaCO3 decreased the a-ketoglutarate formation but increased C(PYR)/ C(alphaKG); and (2) under vitamin limitation conditions increasing the concentration of CaCO3 led to an increased a-KG accumulation at the expenses of pyruvate. To study which ions in CaCO3 was responsible for the accumulation of alpha-KG, the effects of different pH buffers on the a-KG accumulation were studied. The level of alpha-KG was found to correlate with the levels of both Ca2+ and CO3(2-), with Ca2+ played a dominant role and CO3(2-) played a minor role. To find out which pathway was responsible for the accumulation of alpha-KG, the effects of biotin and thiamine on alpha-KG accumulation was investigated. The increase in biotin concentration led to an increase in alpha-KG accumulation and a decrease in C(PYR)/ C(alpha-KG), while the levels of alpha-KG and C(PYR)/C(alphaKG) were not affected by thiamine concentration. The activity of pyruvate carboxylase was increased as much as 40% when the medium was supplemented with Ca2+ . On the other hand, the activity of the pyruvate dehydrogenase complex was unaffected by the presence of Ca2+. To conclude, the higher level of a-KG was caused by higher activity of pyruvate carboxylase stimulated by Ca2+, with CO3(2-) served as the substrate of the reaction.
Liu
Li-Ming
LM
Key Lab of Industrial Biotechnolog o f Ministry of Education, Southern Yangtze University, Wuxi 214036, China.
Li
Yin
Y
Du
Guo-Cheng
GC
Chen
Jian
J
D001709:Biotechnology / Q000379:methods; D002119:Calcium Carbonate / Q000494:pharmacology*; D041221:Candida glabrata / Q000187:drug effects* / Q000378:metabolism*; D005285:Fermentation / Q000187:drug effects*; D007656:Ketoglutaric Acids / Q000378:metabolism*; D008954:Models, Biological; D019289:Pyruvic Acid / Q000378:metabolism*
D007656:Ketoglutaric Acids; D019289:Pyruvic Acid; D002119:Calcium Carbonate