Evaluation of 3-hydroxypropionate biosynthesis in vitro by partial introduction of the 3-hydroxypropionate/4-hydroxybutyrate cycle fromMetallosphaera sedula

Abstract

The chemical 3-hydroxypropionate (3HP) is an important starting reagent for the commercial synthesis of specialty chemicals. In this study, a part of the 3-hydroxypropionate/4-hydroxybutyrate cycle from Metallosphaera sedula was utilized for 3HP production. To study the basic biochemistry of this pathway, an in vitro-reconstituted system was established using acetyl-CoA as the substrate for the kinetic analysis of this system. The results indicated that 3HP formation was sensitive to acetyl-CoA carboxylase and malonyl-CoA reductase, but not malonate semialdehyde reductase. Also, the competition between 3HP formation and fatty acid production was analyzed both in vitro and in vivo. This study has highlighted how metabolic flux is controlled by different catalytic components. We believe that this reconstituted system would be valuable for understanding 3HP biosynthesis pathway and for future engineering studies to enhance 3HP production.Electronic supplementary materialThe online version of this article (doi:10.1007/s10295-016-1793-z) contains supplementary material, which is available to authorized users.

Enzymes, In Vivo Biocatalysis, and Metabolic Engineering for Enabling a Circular Economy and Sustainability

“…However, availability of the key precursor, malonyl-CoA, is generally limited for 3-HP synthesis because the compound is important for cellular synthesis of other important compounds such as ketones and fatty acids . Another challenge of 3-HPA synthesis is the rate-limiting step of this pathway, previously identified as the reaction of Mcr . Effects of Mcr and Msr on 3-HP production were investigated using in vitro assays and measurement of steady-state kinetics parameters of Mcr and Msr.…”

“…704 Another challenge of 3-HPA synthesis is the rate-limiting step of this pathway, previously identified as the reaction of Mcr. 705 HP synthesis from the β-alanine pathway (Figure 61). These engineering efforts yielded a final titer of 665 mg/L 3-HP.…”

Process engineering for microbial production of 3-hydroxypropionic acid

“…Attenuating the latter pathway leads to higher intracellular levels of malonyl-CoA, thus improving 3-HP production. The fatty acid synthesis was primarily inhibited by using cerulenin (Li et al, 2015;Lynch et al, 2011;Rogers and Church, 2016;Ye et al, 2016). Due to the cost of adding such an inhibitor to a fermentation broth, alternative approaches were proposed through genetic engineering (X.…”

“…Due to the cost of adding such an inhibitor to a fermentation broth, alternative approaches were proposed through genetic engineering (X. Lynch et al, 2011;Ye et al, 2016). Although the disruption of genes involved in central metabolism can improve the 3-HP yield, it was also observed that it can hinder cell growth (Ko et al, 2017;Lynch et al, 2011;Tokuyama et al, 2014).…”