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Metabolic fate of glutamate and evaluation of flux through the 4‐aminobutyrate (GABA) shunt in Aspergillus niger
Author(s) -
Kumar Santosh,
Punekar Narayan S.,
SatyaNarayan V.,
Venkatesh K. V.
Publication year - 2000
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/(sici)1097-0290(20000305)67:5<575::aid-bit8>3.0.co;2-l
Subject(s) - glutamate receptor , acidogenesis , biochemistry , chemistry , glutamic acid , metabolism , amino acid , biology , organic chemistry , receptor , methane , anaerobic digestion
Accumulation of GABA and a concurrent block in the Krebs cycle suggest a functional GABA bypass in the acidogenic Aspergillus niger. Apart from the demonstration of enzyme machinery required, a direct measurement of flux through this glutamate decarboxylation loop was attempted. The distribution of carbon from glucose and glutamate was studied using A. niger mycelia grown on different media. The uptake and incorporation of 14 C label into organic acids and amino acids was followed by paper chromatography. Flow of label from glucose into citrate, glutamate and GABA increased in cells harvested at later stages of acidogenic growth. Very little citrate was derived from glutamate while ten times more label reached GABA from labeled glutamate. Radioactivity from L ‐[U‐ 14 C]glutamate and not from L ‐[1‐ 14 C]glutamate was recovered in GABA. This demonstrated that α‐decarboxylation of L ‐glutamate was the source of GABA. Unless grown on GABA, A. niger mycelia did not take up externally supplied GABA. A direct measure of GABA shunt flux was thus not feasible. Therefore a combination of metabolite balance technique and the kinetic approach was applied to evaluate flux from glutamate to succinate in normal and acidogenic A. niger . The flux relative to TCA cycle was estimated by using uptake rate for radiolabeled glutamate, rate of accumulation of certain metabolites and the reactions of GABA metabolism. The analysis indicated that GABA shunt is operative in A. niger and its operation is enhanced during acidogenic growth of the fungus. This is the first report of an estimation of the flux through GABA shunt in a fungus. © 2000 John Wiley & Sons, Inc. Biotechnol Bioeng 67: 575–584, 2000.