Regulatory Mechanism Of BchCXYZ Opera In Rhodopseudomonas Palustris HN1 Strain

Rhodopseudomonas palustris thalli is an important kind of photosynthetic bacteria with rich nutrition. It can use light to produce ATP for growth and has been widely used in aquaculture, wastewater treatment and bio-energy production. As it can use light energy, the study of the expression and regulation mechanism in photosynthetic system is of great importance. The bchCXYZ opera of Rhodopseudomonas palustris not only encodes a series of enzymes catalytic synthesis bacteriochlorophyll(Bch) of the photosystem, but also cotranscribes with its downstream opera pufBALM and therefore plays important roles in photosystem of the bacteria. In the present study, a high hydrogen production strains, HN1, which was isolated in our laboratory previously, was used for the study to clarify the functional mechanisms of bchCXYZ opora in photosystem of Rhodopseudomonas palustris. The major results obtained are as follows:1. We cloned ppsR1 gene from Rhodopseudomonas palustris HN1 strain and found that a frame shift mutation exists in the sequence of ppsR1 gene between Rhodopseudomonas palustris CGA009 and HN1 strains. The DNA binding domain of PpsR protein is highly conserved among many different bacteria strains. We also cloned the promoter region of bchCXYZ gene in Rhodopseudomonas palustris HN1 strain and found a PpsR binding site in the promoter region of bchCXYZ as similar as reported in Bradyrhizobium sp. ORS 278 strain. These results suggested that PpsR1 might also participate in the regulation of bchCXYZ opera in the Rhodopseudomonas palustris HN1 strain.2. Using the bacteria one-hybrid system by constructing two vectors, fusion expressing vector pB1H2ω2-PpsRHTH and reporter vector pH3U3-bchp, the interaction between PpsR1 and bchCXYZ promoter region was investigated. Under the circumstances of the pH3U3-bchp reporter gene self-activiating, two vectors which contain ω2-PpsR and ω2-Zif268 fusion proteins respectively, were individually transformed into the pH3U3-bchp-bait strain. The results obtained showed that the bacteria colony containing theω2-PpsR protein was smaller than that containing theω2-Zif268 protein. Based on the test of ω2-PpsR toxicity and the analysis of reporter gene expression, we confirmed the interation bewteen ω2-PpsR and bchCXYZ promoter region.3. The bacteria one-hybrid system derived from Wolfe lab was employed to screen the regulators of bchCXYZ opera. By modified the fusion expressing vector and constructed a pB1H2ω2-MCS vector, we established a genome library of Rhodopseudomonas palustris HN1. Then, we constructed a series of bait vectors using different promoter regions as the bait to screen the genome library and many potential targeted sequences were obtained.In conclusion, PpsR1 may regulate the bchCXYZ opera in the Rhodopseudomonas palustris HN1 strain. The results obtained from the genome library screening lay a foundation for the discovery of new regulators of bchCXYZ opera in the future.