Additional potential bottlenecks PF299 cost in hydrogen production Biological hydrogen photoproduction is a complex process that requires a tight control/regulation of many pathways at different levels. Genetic engineering has been employed to overcome these limitations and, in most cases, hydrogen production rates have been improved. However, additional genetic modification will be required to achieve maximal conversion efficiency of

solar energy into biohydrogen. These include but are not limited to (a) designing an Gamma-secretase inhibitor inducible leaky ATP synthase mutant and/or inducible proton channel, whereby the proton gradient is dissipated while the cell produces H2; (b) increasing the size of the PQ pool to ameliorate the rate-limiting step in photosynthetic electron transport, buy Bucladesine the oxidation of the PQ pool; and (c) overexpressing NDA2 to increase electron flux into and from the indirect hydrogen production pathway. High-throughput screening techniques To screen for mutants altered in H2 production, several techniques have been developed in the past years as described below. One of the best available methods is a solid-state chemochromic H2 sensor consisting of tungsten oxide and palladium. The palladium captures H2 and transfers it to the tungsten oxide which turns blue when reduced. Chlamydomonas insertional

mutants plated on Petri dishes were screened for attenuated hydrogen production following induction in an anaerobic glove box overnight. When exposed to the light, the cells photoevolved H2, which was detected as blue dots on the H2 sensor (Seibert et al. 2001; Flynn et al. 2002). This method was successfully used to identify the hydrogenase catalytic cluster assembly genes HYDEF and HYDG (Posewitz et al. 2004a) and a starch-less mutant, sta7, in which hydrogenase gene Acetophenone transcription is repressed (Posewitz et al. 2004b). A

water-soluble color indicator has also been used to screen hydrogen-producing microorganisms. This indicator consists of a coloring agent and a water-soluble derivative of Wilkinson’s catalyst [Tris(triphenylphosphine) rhodium chloride]. In this screen, methyl orange and the sulfonate catalyst are dissolved in water and change color when in contact with hydrogen gas. This system can be used with any H2-producing microorganism (Katsuda et al. 2006). Finally, a new and very sensitive technique was recently developed, based on the sensing system from Rhodobacter capsulatus—which acts to upregulate the expression of the native cell’s uptake hydrogenase in response to H2. The Rhodobacter system is composed of the H2-sensor protein (HupUV), a histidine kinase (HupT), a transcription regulator (HupR), and an uptake hydrogenase (HupSL). In the absence of H2, the sensor HupUV interacts with the kinase HupT inducing its autophosphorylation (Elsen et al. 1993).