Two major challenges limit efficient biological H₂ production: inactivation of the hydrogenase enzyme by O₂; and limited electron flow from the photosynthetic apparatus to the hydrogenase.

To address the first challenge, the hypothesis is that the enzyme can be protected by local micro-oxic environments created by nearby O₂ uptake mechanisms. Several complementary ways to reduce O₂ at the vicinity of the hydrogenase will be pursued including use of chimeric proteins in which the hydrogenase is joined with a partner protein capable of reducing O₂ or reactive oxygen species (e.g. glucose oxidase, flavodiiron protein). These chimeras will first be tested in vitro and then the most promising ones will be expressed in vivo.

Rapid molecular and spectroscopic tests will be used to identify limitations to light-driven hydrogen production in the engineered strains. Several genetic modifications will be utilized to rectify identified limitations in electron flow or H₂ production activity.

The complementary sets of expertise in the two research groups in the US and Israel will be put to use in the creation, analysis, and optimization of the engineered algal cells. Together the two groups will determine the optimal way to arrange the various new components to make sustained high-level bio-hydrogen production a reality.

I do not view hydrogen so much as a fuel, but as an essential commodity that we consume at a rate of over 20 million metric tons per year—and which we now make by steam reformation of fossil fuels, a process that is energy intensive and produces carbon dioxide. If we could replace even a part of that with algal biohydrogen that is made via light and water, it would have a substantial impact. However, the state of the biohydrogen field is not even close to where it needs to be in order to be commercially viable.

We thought that some radically different approaches needed to be taken — thus, our crazy idea of hooking up the hydrogenase enzyme directly to Photosystem I in order to divert a large fraction of the electrons from water splitting (by Photosystem II) to make molecular hydrogen.

—Kevin Redding

Plants and algae, as well as cyanobacteria, use photosynthesis to produce oxygen and “fuels,” the latter being oxidizable substances like carbohydrates and hydrogen. There are two pigment-protein complexes that orchestrate the primary reactions of light in oxygenic photosynthesis: Photosystem I (PSI) and Photosystem II (PSII).

Algae (in this case the single-celled green alga Chlamydomonas reinhardtii, or ‘Chlamy’ for short) possess an enzyme called hydrogenase that uses electrons it gets from the protein ferredoxin, which is used to ferry electrons from PSI to various destinations. The algal hydrogenase is rapidly and irreversibly inactivated by oxygen that is constantly produced by PSII. It is hoped that linking the hydrogenase directly to PSI will mitigate the problems, including the fact that hydrogenase competes poorly for electrons and that it is inactivated by oxygen.

In a future commercial system, one will want to be able to grow the cells normally at first, and then switch them to a mode in which most of the electrons are diverted to make hydrogen — essentially crossing over from a cheap replicating system to a “biofactory” in which sunlight drives production of hydrogen using water. The proposed systems provide an obvious way to do that by turning on the genes encoding the linked PSI-hydrogenase proteins. Consequently, electrons will be diverted away from carbon dioxide fixation to hydrogen production.

Iftach is taking very different approaches to this problem, which I see nobody else out there doing. Some of his work is a little controversial, but I think his basic conclusions are sound. We have been talking to each other on and off for a few years, but recently we came to realize that our approaches and skills are very complementary. It is a natural partnership. We are already working on our first two joint manuscripts!

—Kevin Redding

Redding is also partnering with ASU’s Global Institute of Sustainability to develop a module within their Wells Fargo Regional Sustainability Teachers Academy. They are working with Molly Cashion and Robert McGehee, the Academy Program Coordinators.

The team will develop a module on screening algae with an agar overlay method. They will train local middle and high school teachers how to do this in the Academy. They will need only a microwave oven and water bath to perform the assay, and their students will build their illuminators out of a cardboard box using LED strips and AA batteries. Undergraduate student volunteers will bring other materials to classrooms and assist the teachers as needed. Algae are grown on plates, covered with agar mixed with Rhodobacter, and allowed to develop overnight.

The students can image them the next day with their own phone cameras using a small green interference filter provided by the grant. They can then draw their own conclusions about the best hydrogen-producing strains. This plan draws upon concepts from the next-generation science teaching concepts, in which learning is driven by the students’ own curiosity. They will be given only a cursory explanation at first but, as the experiment progresses, the scientists will answer their questions about how things work. The students will be encouraged to experiment with different conditions so as to discover the best algal strains and how to coax them to make more hydrogen. In this way, they become partners in discovery.