Type

Journal Article

Authors

Alexander Steinbüchel
Kevin E O'Connor
Ramesh P Babu
Laura Morales-Gamez
Shane T Kenny
Philipp Fricke
Matthias Raberg
Daniel Heinrich

Subjects

Chemistry

Topics
fatty acid molecular weight gene expression carbon monoxide carbon dioxide amino acid organic waste chain length

Syngas-derived medium-chain-length PHA synthesis in engineered Rhodospirillum rubrum. (2016)

Abstract The purple non-sulfur alphaproteobacterium Rhodospirillum rubrum S1 was genetically engineered to synthesize a heteropolymer of mainly 3-hydroxydecanoic acid and 3-hydroxyoctanoic acid [P(3HD-co-3HO)] from CO- and CO2-containing artificial syngas. For this, genes from Pseudomonas putida KT2440 coding for a 3-hydroxyacyl-ACP thioesterase (phaG), a medium-chain-length (MCL) fatty acid CoA-ligase (PP_0763) and a MCL PHA-synthase (phaC1) were cloned and expressed under the control of the CO-inducible promoter PcooF from R. rubrum S1 in a PHA-negative mutant of R. rubrum P(3HD-co-3HO) was accumulated to up to 7.1 % (wt/wt) of the cell dry weight by a recombinant mutant strain utilizing exclusively the provided gaseous feedstock syngas. In addition to an increased synthesis of these medium chain-length PHAs (PHAMCL), enhanced gene expression through the PcooF promoter led also to an increased molar fraction of 3HO in the synthesized copolymer when compared with the Plac promoter, which regulated expression on the original vector. The recombinant strains were able to partially degrade the polymer, and the deletion of phaZ2, which codes for a PHA-depolymerase most likely involved in intracellular PHA-degradation, did not reduce mobilization of the accumulated polymer significantly. However, an amino acid exchange in the active site of PhaZ2 led to a slight increase in PHAMCL accumulation. The accumulated polymer was isolated; it exhibited an average molecular weight (Mw) of 124.3 kDa and a melting point of 49.6 °C. With the metabolically engineered strains presented in this proof-of-principle study, we demonstrated the synthesis of elastomeric second generation biopolymers from renewable feedstocks not competing with human nutrition. Polyhydroxyalkanoates (PHAs) are natural biodegradable polymers (biopolymers) showing properties similar to commonly produced petroleum-based non-degradable polymers. The utilization of cheap substrates for the microbial production of PHAs is crucial to lower the production costs. Feedstock not competing with human nutrition is highly favorable. Syngas, a mixture of carbon monoxide, carbon dioxide, and hydrogen can be obtained by pyrolysis of organic waste and can be utilized for PHA-synthesis by several bacteria. Up to now, the biosynthesis of PHAs from syngas has been limited to short-chain-length PHAs, which results in a stiff and brittle material. In this study the syngas-utilizing bacterium Rhodospirillum rubrum was genetically modified to synthesize a polymer, which consisted of medium-chain-length constituents resulting in rubber-like material. This study reports the establishment of a microbial synthesis of these so-called medium-chain-length PHAs from syngas and therefore potentially extends the applications of syngas-derived PHAs.
Collections Ireland -> Trinity College Dublin -> PubMed

Full list of authors on original publication

Alexander Steinbüchel, Kevin E O'Connor, Ramesh P Babu, Laura Morales-Gamez, Shane T Kenny, Philipp Fricke, Matthias Raberg, Daniel Heinrich

Experts in our system

1
Kevin E O'Connor
University College Dublin
Total Publications: 52
 
2
Ramesh P Babu
University College Dublin
Total Publications: 11
 
3
Shane T Kenny
University College Dublin
Total Publications: 15