Study On The Mechanism Of Enhanced Photosynthetic Hydrogen Production Ability In NaHSO₃-treated Cyanobacteria Mutant ?135

As a result of increasing awareness regarding environmental problems caused by our present use of fossil fuel-based energy,such as the greenhouse effect,efforts have been undertaken to develop clean and sustainable energy sources.Among many methods that produce clean energy sources,photosynthetic hydrogen production by microalgae is a process by which microalgae uses solar energy and water to produce hydrogen through their own physiological metabolic activities.This method of hydrogen production is clean and pollution-free and has high development potential.At present,many challenges remain to be solved in the photosynthetic hydrogen production by microalgae for industrial requirements.First,microalgae release oxygen during photosynthesis,and hydrogenase is extremely sensitive to oxygen;second,the hydrogen metabolism pathway of microalgae is complex,and hydrogenase requires more abundant electrons that are obtained from photosynthetic electron transport chain.Therefore,it is important for the industrial production of microalgae photosynthetic hydrogen production to explore the anaerobic technology of photosynthetic hydrogen production by microalgae and the key regulatory sites that affect the hydrogen metabolism of microalgae.Based on our previous studies,when the microalgae cells are treated with appropriate concentration of NaHSO₃,this drug can effectively reduce the reactive oxygen species in the cells and activate the expression of hydrogenase.After high-light culture,the microalgal cells can produce superoxide anions and then,reaction of NaHSO₃with superoxide anions indirectly reduces the oxygen content in the cells,thereby activating the activity of hydrogenase and increasing the accumulation levels of photosynthetic hydrogen production by microalgae.After successfully creating an anaerobic environment,the amount of electrons obtained by hydrogenase will directly affect the efficiency of photosynthetic hydrogen production.Clearly,if the PS? activity of microalgae can be maintained at a high level under anaerobic conditions,more sustainable hydrogen production from photosynthesis of microalgae will be achieved.This thesis was carried out under the background of adding NaHSO₃to induce hydrogen production.We selected Synechocystis sp.PCC 6803 as the experimental material because of its clear genetic background.The Synechocystis 6803 transposon mutant library was analyzed by the high-light addition of NaHSO₃to induce hydrogen production.After screening,a mutant strain?135 that grows better than the wild type and has higher PS? activity was obtained,and it was found that the deletion of locus135 of the unknown function gene significantly increased the photosynthetic hydrogen production of Synechocystis 6803.We found that under high light,?135 has a phenotype that grows slower and more likely to turn yellow than wild type,but the activity of PS? is better than that of wild type.Under high light,the PSI activity of?135 was damaged.At this time,the cells of?135 produced more superoxide anions than the wild type.After adding a certain concentration of NaHSO₃to the algae solution,within 12 hours,the PSI activity of?135 recovered to a level similar to that of the wild type;after adding NaHSO₃,the dissolved oxygen in the?135 cells drops to zero more quickly and enters an anaerobic state,thereby retaining more PS? activity,and achieving a longer duration of photosynthetic hydrogen production.As a consequence,the 36-hour hydrogen production accumulation of the mutant strain is about 1.6 times than that of the wild type.By measuring the levels of hydrogen production in the presence of PS? inhibitor lincomycin(Lin),carbon assimilation inhibitor glycolaldehyde(GA)and PSI cycle electron transport inhibitor antimycin A(AA)or not,we found that the electron source of photosynthetic hydrogen production is stem from the PS? photolysis water.The fundamental reason why?135 mutant can produce higher hydrogen than wild-type is because it has higher PS? activity that can provide a more abundant electron source for hydrogenase.This thesis explores the reasons for the high hydrogen production of the mutant?135 under the background of NaHSO₃,hoping to provide a reference for the future industrial application of microalgae photosynthetic hydrogen production.