Читать книгу Biomolecular Engineering Solutions for Renewable Specialty Chemicals - Группа авторов - Страница 55

Biohydrogen is the clean fuel as it burns leaving only water. It is produced biologically and therefore is a promising future fuel (Hosseinpour et al., 2017). Hydrogenase or nitrogenase are the crucial enzyme for biohydrogen regulation in both prokaryotes and eukaryotes. Autotrophic organisms such as microalgae, green algae, cyanobacteria, etc. are most efficient for biohydrogen production. The main principle behind the hydrogen production is the electrons generated during metabolism inside the cells by the action of hydrogenase enzyme are made to form hydrogen. Genetic manipulation and metabolic engineering of autotrophic organisms for production of biohydrogen has received much attention.

In cyanobacteria three alternative pathways (i) photolysis of water using photosystems (PS); (ii) fermentative pathway; and (iii) photofermentative pathway for the production of biohydrogen. In the first pathway, PSI and PSII through light‐dependent reaction transfer electron from water to ferredoxin producing NADPH leading to biohydrogen formation (Carrieri et al., 2011). In second, the source of NADPH is degraded polysaccharides or lipids, and the electrons are transferred to plastoquinone pool for hydrogen formation (Ghirardi et al., 2000). Third pathway is the combination of first two. Cyanobacteria having heterocyst use it as the sites for nitrogen fixation. A vegetative cell originates NADPH and transport electrons to the plastoquinone pool inside the heterocyst. Inside this hydrogenase gets inactivated and nitrogenase reaction take place (Kufryk, 2013). Before the hydrogen production by cyanobacteria can become industrially viable improvements in strains are to be done. There are two types of hydrogenase: uptake hydrogenase (Hup) and bidirectional hydrogenase (Hox). Anabaena sp. PCC 7120 contains both Hup and Hox (Masukawa et al., 2002). The species were tested for both Hup and Hox inactivation. Hup inactivated strain gave four to seven times increase in H₂ production. In another study, Nostoc sp. PCC 7422 was chosen having highest nitrogenase activity and was subjected to Hup gene disruption (Yoshino et al., 2007). The generated mutant was able to produce hydrogen at a rate of threefold than wild type. Electrons play an important role in hydrogen production. Electrons generated by ferredoxin and NADPH are sometimes transferred to other competing pathways (e.g. nitrate assimilation pathway). This can be engineered to redirect electron flow to, for example, Hox (Baebprasert et al., 2011). Disrupting nitrate assimilating pathway in Synechocystis sp. strain PCC 6803 results in higher hydrogen production as electron gets redirected to Hox. LDH mutants of Synechococcus 7002 lacking have exhibited 5 times greater hydrogen production compared with the wild type (Park et al., 2008). In a different study, Cyanothece sp. ATCC 51142, a unicellular diazotrophic cyanobacterium demonstrated the ability to generate high levels of hydrogen (465 μmol H₂/mg of chlorophyll h) using glycerol as a substrate under aerobic conditions (Bandyopadhyay et al., 2010).

Green algae have two light‐dependent and one light‐independent pathway mediated by [Fe] or [FeFe] hydrogenase for hydrogen synthesis (Meyer, 2007). In it [Fe] or [FeFe] hydrogenase is the key catalyst and ferredoxin is electron donor. In the first pathway, water is used as the electron sink and biophotolysis of water takes place. Here water splitting PSII and ferredoxin‐reducing PSI act together. The second pathway is PSII independent where electrons from metabolic pathways like glycolysis or citric acid are transferred to electron transport chain. The third pathway that is light independent or dark fermentation of decarboxylated pyruvate. Chlorella vulgaris strain when overexpressed with hydrogenase gene can produce hydrogen even in atmospheric conditions (Hwang et al., 2014). These strains were able to show maximum production at low intensities of light much less than that is received on Earth by Sun. This can be overcome by decreasing the number of light harvesting antenna as done in Chlamydomonas reinhardtii tla1 strain (Kosourov et al., 2011). In total, 50% truncated photosynthetic light harvesting antenna increases hydrogen production. Flavodiiron (Flv3B) proteins have crucial and specific roles in photoprotection of photosystems I and II in cyanobacteria. Absence of Flv3B leads to impaired growth of cyanobacteria in the presence of oxygen. Overexpression of Flv3B in Nostoc PCC7120 significantly increases the hydrogen production (Roumezi et al., 2020).