Biology subjects

Rengasamy, K.

Publications and source records attributed to Rengasamy, K..

5 recordsLinked to original sources

bioRxiv2023

Improving bioplastic production by Rhodopseudomonas palustris TIE-1 using synthetic biology and metabolic engineering

With the increasing demand for sustainably produced renewable resources, it is important to look towards microorganisms capable of producing bioproducts such as biofuels and bioplastics. Though many systems for bioproduct production are well documented and tested in model organisms, it is essential to look beyond to non-model organisms to expand the field and take advantage of metabolically versatile strains. This investigation centers on Rhodopseudomonas palustris TIE-1, a purple, non-sulfur autotrophic, and anaerobic bacterium capable of producing bioproducts that are comparable to their petroleum-based counterparts. To induce bioplastic overproduction, genes that might have a potential role in the PHB biosynthesis such as the regulator, phaR, and phaZ known for its ability to degrade PHB granules were deleted using markerless deletion. Mutants in pathways that might compete with polyhydroxybutyrate (PHB) production such as glycogen and nitrogen fixation previously created to increase n-butanol production by TIE-1 were also tested. In addition, a phage integration system was developed to insert RuBisCO (RuBisCO form I and II genes) driven by a constitutive promoter PaphII into TIE- 1 genome. Our results show that deletion of the phaR gene of the PHB pathway increases PHB productivity when TIE-1 was grown photoheterotrophically with butyrate and ammonium chloride (NH4Cl). Mutants unable to make glycogen or fix dinitrogen gas show an increase in PHB productivity under photoautotrophic growth conditions with hydrogen. In addition, the engineered TIE-1 overexpressing RuBisCO form I and form II produces significantly more polyhydroxybutyrate than the wild type under photoheterotrophy with butyrate and photoautotrophy with hydrogen. Inserting RuBisCO genes into TIE-1 genome is a more effective strategy than deleting competitive pathways to increase PHB production in TIE-1. The phage integration system developed for TIE-1 thus creates numerous opportunities for synthetic biology in TIE-1.

microbiology↗

bioRxiv2023

The phototrophic bacteria Rhodomicrobium spp. are novel chassis for bioplastic production

Polyhydroxybutyrate (PHB) is a bio-based, biodegradable alternative to petroleum-based plastics. PHB production at industrial scales remains infeasible, in part due to insufficient yields and high costs. Addressing these challenges requires identifying novel biological chassis for PHB production and modifying known biological chassis to enhance production using sustainable, renewable inputs. Here, we take the former approach and present the first description of PHB production by two prosthecate photosynthetic purple non-sulfur bacteria (PNSB), Rhodomicrobium vannielii and Rhodomicrobium udaipurense. We show that both species produce PHB across photoheterotrophic, photoautotrophic, photoferrotrophic, and photoelectrotrophic growth conditions. Both species show the greatest PHB titers during photoheterotrophic growth on butyrate with dinitrogen gas as a nitrogen source (up to 44.08 mg/L), while photoelectrotrophic growth demonstrated the lowest titers (up to 0.13 mg/L). These titers are both greater (photoheterotrophy) and less (photoelectrotrophy) than those observed previously in a related PNSB, Rhodopseudomonas palustris TIE-1. On the other hand, we observe the highest electron yields during photoautotrophic growth with hydrogen gas or ferrous iron electron donors, and these electron yields were generally greater than those observed previously in TIE-1. These data suggest that non model organisms like Rhodomicrobium should be explored for sustainable PHB production and highlights utility in exploring novel biological chassis.

microbiology↗

bioRxiv2020

Sustainable Production of the Biofuel n-Butanol by Rhodopseudomonas palustris TIE-1

Anthropogenic carbon dioxide (CO2) release in the atmosphere from fossil fuel combustion has inspired scientists to study CO2 to fuel conversion. Oxygenic phototrophs such as cyanobacteria have been used to produce biofuels using CO2. However, oxygen generation during oxygenic photosynthesis affects biofuel production efficiency. To produce n-butanol (biofuel) from CO2, here we introduced an n-butanol biosynthesis pathway into an anoxygenic (non-oxygen evolving) photoautotroph, Rhodopseudomonas palustris TIE-1 (TIE-1). Using different carbon, nitrogen, and electron sources, we achieved n-butanol production in wild-type TIE-1 and mutants lacking electron-consuming (nitrogen-fixing) or acetyl-CoA-consuming (polyhydroxybutyrate and glycogen synthesis) pathways. The mutant lacking the nitrogen-fixing pathway produced highest n-butanol. Coupled with novel hybrid bioelectrochemical platforms, this mutant produced n-butanol using CO2, solar panel-generated electricity, and light, with high electrical energy conversion efficiency. Overall, this approach showcases TIE-1 as an attractive microbial chassis for carbon-neutral n-butanol bioproduction using sustainable, renewable, and abundant resources.

bioengineering↗

bioRxiv2017

Improving microbial electrosynthesis of polyhydroxybutyrate (PHB) from CO2 by Rhodopseudomonas palustris TIE-1 using an immobilized iron complex modified cathode

Microbial electrosynthesis (MES) is a promising bioelectrochemical approach to produce biochemicals. A previous study showed that Rhodopseudomonas palustris TIE-1 can directly use poised electrodes as electron donors for photoautotrophic growth at cathodic potentials that avoid electrolytic H2 production (photoelectroautotrophy). To make TIE-1 an effective biocatalyst for MES, we need to improve its electron uptake ability and growth under photoelectroautotrophic conditions. Because TIE-1 interacts with various forms of iron while using it as a source of electrons for photoautotrophy (photoferrotrophy), we tested the ability of iron-based redox mediators to enhance direct electron uptake. Our data show that soluble iron cannot act as a redox mediator for electron uptake by TIE-1 from a cathode poised at +100mV vs. Standard Hydrogen electrode. We then tested whether an immobilized iron-based redox mediator Prussian Blue (PB) can enhance electron uptake by TIE-1. Chronoamperometry indicates that cathodic current uptake by TIE-1 increased from 1.47 {+/-} 0.04 to 5.6 {+/-} 0.09 {micro}A/cm2 (3.8 times) and the production of the bioplastic, polyhydroxybutyrate (PHB) improved from 13.5 {+/-} 0.2 g/L to 18.8 {+/-} 0.5 g/L (1.4 times) on electrodes coated with PB. Overall, our data show that immobilized PB can increase direct electron uptake by TIE-1 and enhances PHB production.

microbiology↗

bioRxiv2017

Towards sustainable bioplastic production in resource limited environments using the photoferroautotrophic and photoelectroautotrophic bacterium Rhodopseudomonas palustris TIE-1

Bioplastics are an attractive alternative to petroleum-derived plastics because of the harmful environmental effects of conventional plastics and the impending fossil fuel crisis. Polyhydroxybutyrate (PHB) is a well-known bioplastic that is produced by several microbes using organic carbon sources. Autotrophic (using carbon dioxide or CO2) PHB production is reported for only a few organisms. Sustainable PHB bioproduction using other autotrophic microbes needs to be explored. Rhodopseudomonas palustris, a metabolically versatile purple non-sulfur bacterium (PNSB) is known to produce PHBs under photoheterotrophic conditions. Rhodopseudomonas palustris strain TIE-1 demonstrates extended metabolic versatility by using electron sources such as ferrous iron and poised electrodes for photoautotrophy. Here we report the ability of TIE-1 to produce PHB under photoferroautotrophic (light - energy source, ferrous iron - electron source and CO2 - carbon source) and photoelectroautotrophic (light - energy source, poised electrodes - electron source and CO2 - carbon source) growth conditions. PHB accumulation was observed both under nitrogen (N2) fixing and non-N2 fixing conditions. For comparison, we determined PHB production under chemoheterotrophic, photoheterotrophic and photoautotrophic conditions using hydrogen as the electron donor. Photoferroautotrophic and photoelectroautotrophic PHB production was on par with that observed from organic carbon substrates such as butyrate. PHB production increased during N2 fixation under photoheterotrophic conditions but not during photoautotrophic growth. Electron microscopy confirmed that TIE-1 cells accumulate PHBs internally under the conditions that showed highest production. However, gene expression analysis suggests that PHB cycle genes are not differentially regulated despite observable changes in biopolymer production.

microbiology↗