Namita Khanna

Cyanobacterial Hydrogenases and Hydrogen Metabolism Revisited: Recent Progress and Future Prospects

Cyanobacteria have garnered interest as potential cell factories for hydrogen production. In conjunction with photosynthesis, these organisms can utilize inexpensive inorganic substrates and solar energy for simultaneous biosynthesis and hydrogen evolution. However, the hydrogen yield associated with these organisms remains far too low to compete with the existing chemical processes. Our limited understanding of the cellular hydrogen production pathway is a primary setback in the potential scale-up of this process. In this regard, the present review discusses the recent insight around ferredoxin/flavodoxin as the likely electron donor to the bidirectional Hox hydrogenase instead of the generally accepted NAD(P)H. This may have far reaching implications in powering solar driven hydrogen production. However, it is evident that a successful hydrogen-producing candidate would likely integrate enzymatic traits from different species. Engineering the [NiFe] hydrogenases for optimal catalytic efficiency or expression of a high turnover [FeFe] hydrogenase in these photo-autotrophs may facilitate the development of strains to reach target levels of biohydrogen production in cyanobacteria. The fundamental advancements achieved in these fields are also summarized in this review.

Biohydrogen Production: Fundamentals and Technology Advances

This book compiles the fundamentals of biohydrogen production technology. It offers comprehensive coverage of microbiology, biochemistry, feedstock requirements, and molecular biology of the biological hydrogen production processes. It also gives insight into scale-up problems and limitations. In addition, the book discusses mathematical modeling of the various processes involved in biohydrogen production and the software required to model the processes. The book summarizes research advances that have been made in this field and discusses bottlenecks of the various processes, which presently limit the commercialization of this technology.

Turning around the electron flow in an uptake hydrogenase. EPR spectroscopy and in vivo activity of a designed mutant in HupSL from Nostoc punctiforme

The filamentous cyanobacterium Nostoc punctiforme ATCC 29133 produces hydrogen via nitrogenase in heterocysts upon onset of nitrogen-fixing conditions. N. punctiforme expresses concomitantly the uptake hydrogenase HupSL, which oxidizes hydrogen in an effort to recover some of the reducing power used up by nitrogenase. Eliminating uptake activity has been employed as a strategy for net hydrogen production in N. punctiforme (Lindberg et al., Int. J. Hydrogen Energy, 2002, 27, 1291–1296). However, nitrogenase activity wanes within a few days. In the present work, we modify the proximal iron-sulfur cluster in the hydrogenase small subunit HupS by introducing the designed mutation C12P in the fusion protein f-HupS for expression in E. coli (Raleiras et al., J. Biol. Chem., 2013, 288, 18345–18352), and in the full HupSL enzyme for expression in N. punctiforme. C12P f-HupS was investigated by EPR spectroscopy and found to form a new paramagnetic species at the proximal cluster site consistent with a [4Fe–4S] to [3Fe–4S] cluster conversion. The new cluster has the features of an unprecedented mixed-coordination [3Fe–4S] metal center. The mutation was found to produce stable protein in vitro, in silico and in vivo. When C12P HupSL was expressed in N. punctiforme, the strain had a consistently higher hydrogen production than the background ΔhupSL mutant. We conclude that the increase in hydrogen production is due to the modification of the proximal iron-sulfur cluster in HupS, leading to a turn of the electron flow in the enzyme.

Metabolic and genetic engineering of cyanobacteria for enhanced hydrogen production

There is an urgent need to develop sustainable solutions to convert solar energy into energy carriers used in society. In addition to solar cells generating electricity, there are several options to generate solar fuels, with molecular hydrogen being an interesting and promising option. Native and engineered cyanobacteria have been used as model systems to examine, develop and demonstrate photobiological hydrogen production. In the present review we present and discuss recent progress with respect to native biological systems to generate hydrogen, metabolic modulations, and genetic engineering of metabolic pathways, as well as the introduction of custom-designed, non-native enzymes and complexes for enhanced hydrogen production in cyanobacteria. In conclusion, metabolic and genetic engineering of native cyanobacterial hydrogen metabolism can significantly increase the hydrogen production. Introduction of custom-designed non-native capacities open up new possibilities to further enhance cyanobacterial-based hydrogen production.

In vivo activation of an [FeFe] hydrogenase using synthetic cofactors

[FeFe] hydrogenases catalyze the reduction of protons, and oxidation of hydrogen gas, with remarkable efficiency. The reaction occurs at the H-cluster, which contains an organometallic [2Fe] subsite. The unique nature of the [2Fe] subsite makes it dependent on a specific set of maturation enzymes for its biosynthesis and incorporation into the apo-enzyme. Herein we report on how this can be circumvented, and the apo-enzyme activated in vivo by synthetic active site analogues taken up by the living cell.

Engineering Cyanobacteria for Biofuel Production

Fast depletion of petroleum resources and environmental concerns due to rapidly increasing fossil fuel related CO2 emissions have prompted scientists to find more sustainable and environmental friendly fuel alternatives. Considering this, algal conversion of CO2 to biofuels has received increased attention in recent times. In particular, cyanobacteria have been considered as promising candidates for biofuel production considering their fast growth rate, ability to fix carbon dioxide, and their genetic tractability. In parallel to the advancements in synthetic biology and genetic engineering, several proofs of concept studies have emerged demonstrating the ability of cyanobacteria to produce different kinds of biofuels including alcohols, hydrogen, and fatty acid derived biofuels. However, presently their low titer values impede their commercial success. In this perspective, we review the recent publications on engineering cyanobacteria for biofuel production discuss the challenges and scope of improvements for advancing cyanobacterial fuel production.

Fundamentals and Recent Advances in Hydrogen Production and Nitrogen Fixation in Cyanobacteria

There is an urgent need to develop sustainable solutions to convert solar energy into energy carriers used in the society. In addition to solar cells generating electricity, there are several options to generate solar fuels. Native and engineered cyanobacteria have been as model systems to examine, demonstrate, and develop photobiological hydrogen production. In the present contribution the knowledge and understanding of the native systems in cyanobacteria to generate hydrogen, as well as metabolic modulations and genetic engineering to enhance hydrogen production is presented and summarized. Specifically, the recent insight around ferredoxin/flavodoxin as the likely electron donor to the bidirectional Hox-hydrogenase instead of the generally accepted NAD(P)H is highlighted and discussed. In addition, engineering approaches of [NiFe] hydrogenases for optimal catalytic efficiencies and attempts to express high turnover [FeFe] hydrogenase in cyanobacteria that may facilitate the development of strains to reach target levels of hydrogen production in cyanobacteria are detailed. The fundamental advancements achieved in these fields are summarized in this review.

Correction: In vivo activation of an [FeFe] hydrogenase using synthetic cofactors

Correction for ‘In vivo activation of an [FeFe] hydrogenase using synthetic cofactors’ by N. Khanna et al., Energy Environ. Sci., 2017, 10, 1563–1567.

Perspectives on Algal Engineering for Enhanced Biofuel Production

Algae as photoautotrophs can trap the solar energy and convert it into usable form. Solar energy is the most abundant and ultimate energy source. The total amount of solar energy absorbed by the Ea ...