Heterologous ectoine production in Escherichia coli: by-passing the metabolic bottle-neck.

Abstract

Transcription of the ectoine biosynthesis genes ectA, ectB and ectC from Marinococcus halophilus in recombinant Escherichia coli DH5alpha is probably initiated from three individual sigma70/sigmaA-dependent promoter sequences, upstream of each gene. Consequently, mRNA-fragments containing the single genes and combinations of the genes ectA and ectB or ectB and ectC, respectively, could be detected by Northern blot analysis. Under the control of its own regulatory promoter region (ectUp) a seemingly osmoregulated ectoine production was observed. In addition, aspartate kinases were identified as the main limiting factor for ectoine production in recombinant E. coli DH5alpha. Co-expression of the ectoine biosynthesis genes and of the gene of the feedback-resistant aspartate kinase from Corynebacterium glutamicum MH20-22B (lysC) led to markedly increased production of ectoine in E. coli DH5alpha, resulting in cytoplasmic ectoine concentrations comparable to those reached via ectoine accumulation from the medium.