Identification and characterization of the propanediol utilization protein PduP of Lactobacillus reuteri for 3-hydroxypropionic acid production from glycerol

Abstract

Although the de novo biosynthetic mechanism of 3-hydroxypropionic acid (3-HP) in glycerol-fermenting microorganisms is still unclear, the propanediol utilization protein (PduP) of Lactobacillus species has been suggested to be a key enzyme in this regard. To verify this hypothesis, a pduP gene from Lactobacillus reuteri was cloned and expressed, and the encoded protein was characterized. Recombinant L. reuteri PduP exhibited broad substrate specificity including 3-hydroxypropionaldehyde and utilized both NAD(+) and NADP(+) as a cofactor. Among various aldehyde substrates tested, the specific activity was highest for propionaldehyde, at pH 7.8 and 37 °C. The K(m) and V(max) values for propionaldehyde in the presence of NAD(+) were 1.18 mM and 0.35 U mg⁻¹, respectively. When L. reuteri pduP was overexpressed in Klebsiella pneumoniae, 3-HP production remarkably increased as compared to the wild-type strain (from 0.18 g L⁻¹ to 0.72 g L⁻¹) under shake-flask culture conditions, and the highest titer (1.38 g L⁻¹ 3-HP) was produced by the recombinant strain under batch fermentation conditions in a bioreactor. This is the first report stating the enzymatic properties of PduP protein and the probable role in biosynthesis of 3-HP in glycerol fermentation.

Rhodobacter sphaeroides Uses a Reductive Route via Propionyl Coenzyme A To Assimilate 3-Hydroxypropionate

“…The primarily marine substrate DMSP is by far not the only source of 3-hydroxypropionate in nature, but instead, 3-hydroxypropionate is the product or intermediate of the breakdown of many substrates in a variety of environments (4,18,29,30,31,33,41). It will therefore not be surprising to also find many nonmarine organisms that will be able to derive carbon from 3-hydroxypropionate.…”

“…Yet another metabolic process involving 3-hydroxypropionate is the anaerobic metabolism of glycerol. There have also been reports of 3-hydroxypropionate formation by the fermentation of glycerol by lactic acid bacteria (31,41) and the anaerobic oxidation of glycerol by a sulfate-reducing bacterium (33). In summary, 3-hydroxypropionate is likely to play an important role in the overall carbon cycle as an end product or intermediate in the carbon metabolism of a variety of compounds.…”

Diverse Bacterial Microcompartment Organelles

“…Recent studies showed that a short N-terminal sequence targets PduP to the interior of the MCP, as described above (40). Homologs of PduP from other organisms have been purified and studied biochemically (116,117). Some PduP homologs show broad enzymatic activity on various aliphatic aldehydes, including 3-hydroxypropionaldehyde (HPA), as evident from studies with the Klebsiella pneumoniae and Lactobacillus reuteri enzymes (116,117).…”

“…Homologs of PduP from other organisms have been purified and studied biochemically (116,117). Some PduP homologs show broad enzymatic activity on various aliphatic aldehydes, including 3-hydroxypropionaldehyde (HPA), as evident from studies with the Klebsiella pneumoniae and Lactobacillus reuteri enzymes (116,117). These homologs, which share ϳ86% amino acid sequence identity with the Salmonella PduP enzyme, are used for production of 3-hydroxypropionic acid during glycerol degradation (116,117).…”

Heterologous Expression of Aldehyde Dehydrogenase from Saccharomyces cerevisiae in Klebsiella pneumoniae for 3-Hydroxypropionic Acid Production from Glycerol

“…3-HP serves as the versatile precursor of several commercially important chemicals, such as 1,3-propanediol, acrylic acid, acrylamide, acrolein [7,17], polymer [1], or as a potential nematicide [14]. Glycerol-based 3-HP biosynthesis through microbial metabolic engineering has been centered in recent years due to ample glycerol as a main by-product in the flourishing biodiesel industry [2,10].…”