Fig. 1. Metabolic engineering of H. bluephagenesis for 3HP production.

a Overall strategies for 3HP production involving a combination of marker-free dddA deletion and plasmid-expressing dhaT_Pp, aldD_Pp. Red color “X” indicates inactivation of metabolic pathways. Gene dhaT_Pp encodes P. putida KT2440 alcohol dehydrogenase, aldD_Pp encodes P. putida KT2440 aldehyde dehydrogenase, adhP encodes H. bluephagenesis alcohol dehydrogenase, aldD_Hb encodes H. bluephagenesis aldehyde dehydrogenase. DddA putative 3-hydroxypropionate dehydrogenase in H. bluephagenesis, DddC CoA-acylating methylmalonate-semialdehyde dehydrogenase in H. bluephagenesis, PhaA 3-ketothiolase, PhaB acetoacetyl-CoA reductase, PhaC PHA synthase. b 3HP production by E. coli and H. bluephagenesis harboring plasmid p30, respectively. c PHB production by H. bluephagenesis cultured on 5 g L⁻¹ 3HP as a sole carbon source. d Influence of 3HP on cell metabolism at the transcriptional level in H. bluephagenesis. Volcano plot and fold change of gene expression of differentially expressed gene (DEG) distribution. A corrected P-value of 0.05 and log2 (fold change) of 1 were set as the thresholds for significantly differential expression. Red, upregulated genes; blue, downregulated genes. qRT-PCR analysis of several putative upregulated dehydrogenases expression levels in the 3HP containing cultures compared to that of the glucose-containing ones. e 3HP and PHB production by H. bluephagenesis TDΔdddA harboring plasmid p30. Cells were grown in a defined minimal medium supplemented with 30 g L⁻¹ glucose and 10 g L⁻¹ 1,3-propanediol. All titers were obtained after 48 h cultivation at 200 r.p.m. at 37 °C. The initial pH of all shake-flask studies was 9. All data represent the mean of n = 3 biologically independent samples and error bars show s.d. Two-tailed Student’s t-tests were performed to determine the statistical significance for two-group comparisons. DCM dry cell mass, PHB polyhydroxybutyrate.