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. 2004 May;70(5):2861-6.
doi: 10.1128/AEM.70.5.2861-2866.2004.

Heterologous expression of lactose- and galactose-utilizing pathways from lactic acid bacteria in Corynebacterium glutamicum for production of lysine in whey

Eoin Barrett  1 , Catherine Stanton, Oskar Zelder, Gerald Fitzgerald, R Paul Ross
Affiliations

Heterologous expression of lactose- and galactose-utilizing pathways from lactic acid bacteria in Corynebacterium glutamicum for production of lysine in whey

Eoin Barrett et al. Appl Environ Microbiol. 2004 May.

Abstract

The genetic determinants for lactose utilization from Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842 and galactose utilization from Lactococcus lactis subsp. cremoris MG 1363 were heterologously expressed in the lysine-overproducing strain Corynebacterium glutamicum ATCC 21253. The C. glutamicum strains expressing the lactose permease and beta-galactosidase genes of L. delbrueckii subsp. bulgaricus exhibited beta-galactosidase activity in excess of 1000 Miller units/ml of cells and were able to grow in medium in which lactose was the sole carbon source. Similarly, C. glutamicum strains containing the lactococcal aldose-1-epimerase, galactokinase, UDP-glucose-1-P-uridylyltransferase, and UDP-galactose-4-epimerase genes in association with the lactose permease and beta-galactosidase genes exhibited beta-galactosidase levels in excess of 730 Miller units/ml of cells and were able to grow in medium in which galactose was the sole carbon source. When grown in whey-based medium, the engineered C. glutamicum strain produced lysine at concentrations of up to 2 mg/ml, which represented a 10-fold increase over the results obtained with the lactose- and galactose-negative control, C. glutamicum 21253. Despite their increased catabolic flexibility, however, the modified corynebacteria exhibited slower growth rates and plasmid instability.

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Figures

FIG. 1.
FIG. 1.
Schematic diagram of the cloning of the lactose and galactose operons into pCLIK 5a MCS Pddh. pEB01 was constructed by cloning the lactose operon of L. delbrueckii subsp. bulgaricus into the plasmid pCLIK 5a MCS Pddh downstream of the promoter ddH, while pEB02 was constructed by cloning the galactose operon of L. lactis subsp. cremoris MG 1363 into the plasmid pEB01.
FIG. 2.
FIG. 2.
Comparison of the levels of growth of C. glutamicum 21253 (pEB02) in 2% (wt/vol) glucose (▴), lactose (▪), and galactose (•) MM and without a carbon source (⧫) with the growth of C. glutamicum 21253 (pCLIK 5a MCS Pddh) (—) in 2% (wt/vol) glucose.
FIG. 3.
FIG. 3.
Comparison of the levels of growth of C. glutamicum 21253 (pCLIK MCS 5a Pddh) (▴) and C. glutamicum 21253 (pEB02) (×) in whey-based medium (I1). Levels of lysine production by C. glutamicum 21253 (pCLIK MCS 5a Pddh) (▪) and C. glutamicum 21253 (pEB02) (□) in whey-based media (I1) are shown.
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References

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