Strain improvement of Rhizopus oryzae for over-production of l(+)-lactic acid and metabolic flux analysis of mutants

Strain improvement has been conventionally achieved through mutation and selection. However, the genetic and metabolic profiles of mutant strains were poorly characterized and mutagenesis remained a random process. Metabolic flux analysis (MFA) of mutants will enable us to better understand the chan...

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Bibliographic Details
Published in:Biochemical engineering journal 2004-04, Vol.18 (1), p.41-48
Main Authors: Bai, Dong-Mei, Zhao, Xue-Ming, Li, Xin-Gang, Xu, Shi-Min
Format: Article
Language:English
Subjects:
Biological and medical sciences
Biotechnology
Fermentation
Filamentous fungi
Fundamental and applied biological sciences. Psychology
Lactic acid
Metabolic flux analysis
Starch
Strain improvement
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Summary:Strain improvement has been conventionally achieved through mutation and selection. However, the genetic and metabolic profiles of mutant strains were poorly characterized and mutagenesis remained a random process. Metabolic flux analysis (MFA) of mutants will enable us to better understand the change of flux profile in vivo and the mechanism of mutagenesis. A l(+)-lactic acid over-producing mutant, Rhizopus oryzae R1021, was isolated by mutagenizing the parent strain ( R. oryzae R3017) with UV, diethyl sulfate (DES) and 60 Co . Starting with a concentration of 120 g/l corn starch, mutant R1021 produced 79.4 g/l l(+)-lactic acid after 60 h in flasks, 52% higher than that produced by the parent strain. The l(+)-lactic acid purity was 99.05% by weight based on the amount of total lactic acid. The mutant R1021 was also morphologically different from the parent strain. The results of carbon flux analysis of the parent strain and mutants showed that the pyruvate node in the metabolic model was the principle and flexible node. The results show that the key steps of the pathways in parent strain where most carbon is lost from lactic acid formation are the reactions form pyruvate to acetyl-CoA, oxaloacetate and ethanol. Although the fractions of the carbon from pyruvate to ethanol and acetyl-CoA were reduced in mutants, these two pathways are still the steps that are most likely to be further targeted to reroute the pyruvate metabolism to improve lactic acid production.
ISSN:1369-703X
1873-295X
DOI:10.1016/S1369-703X(03)00126-8