Studies On Anabaena Sp.PCC 7120 Gas Vesicle Gene Cluster And Its Coding Proteins

Floattaion at a proper depth in the water and mainintaing the efficient supply of phosphate are two key factors for the survival and growth of cyanobacteria.We have investigated the gas vesicle gene cluster of Anabaena sp.PCC 7120 and the phospahate transport system of marine cyanobacteria,respectively;thus this study includes the following two parts.Part 1:The gas vesicle gene cluster of Anabaena sp.PCC 7120 and its encoded proteinsDifferent modes of bacterial taxis play important roles in environmental adaptation,survival,colonization.One mode of taxis is floatation thanks to the production of gas vesicles.Gas vesicles,which enable the bacteria to float in aquatic niches,are proteinaceous intracellular organelles,permeable only to gas.Gas vesicles are closely related to the growth and survival of prokaryotes by providing buoyancy.Most dominant cyanobacteria that form blooms produce gas vesicles.Harmful bloom caused by cyanobacteria become a major water environmental problem in China's freshwater ecosystems such as Dianchi lake,Taihu lake and Chaohu lake for a long time.Although many strains of cyanobacteria have been reported to be capable of producing gas vesicles in response to various environmental stimili,it is debatable whether gas vesicles are present in Anabaena sp.PCC 7120.In this study,we tested whether Anabaena sp.PCC 7120 is capable of producing gas vesicles at both the transcriptional and translational levels.The gas vesicle gene cluster of Anabaena sp.PCC 7120 contains an intact gvp gene cluster,gvpA1A2CNJKFGVW,and real-time quantitative PCR detection showed that gvpA is constitutively expressed,and could be upregulated upon stimuli such as low temperature and/or high light.The enzymatic activity assays showed that GvpN is an ATPase,and we determined the enzymatic kinetic parameters of GvpN towards ATP.In addition,the structrure of GvpF was sovled at the resolution of 2.5 A,revealing an overall structure highly similar to that of Microcystis PCC7806.Heterologous expression of the intact Anabaena sp.PCC 7120 gvp gene cluster enabled E.coli cells to gain the capacity of floatation thanks to the formation of irregular gas vesicles.Combined with the results of enzymatic and structural studies,these findings revealed that Anabaena sp.PCC 7120 has the ability to form gas vesicles under certain environmental conditions.In addition,gas vesicles are not only important in the regulation of water blooms,but also possess potential application value in medicine as a kind of proteinaceous nanoparticles,which are non-toxic,stable,and of good bioengineering performance and biocompatibility.Gas vesicles can scatter sound waves and are used as contrast agents for contrast-enhanced ultrasound,and have a promising application prospect in magnetic resonance imaging(MRI)due to the gradient microscale magnetic field generated by the gas contained inside and the interaction with the hyperpolarized nuclei.Gas vesicles of Anabaena FACHB 1091 were purified and have been shown to have an enhanced effect on intrahepatic contrast-enhanced ultrasound in rat.Part 2:The phosphate binding proteins of marine cyanobacteria and cyanophagesThe uptake of phosphorus plays an essential role in cell growth.In cyanobacteria,it mainly relies on ABC-type phosphate-specific transport(Pst)system.This ABC-type phosphate transporter comprises a periplasmic high-affinity phosphate-binding protein PstS,a channel formed by two transmembrane proteins PstC and PstA,and the intracellular ATPase PstB.The gene encoding PstS exits in not only cyanobacterial genome but also cyanophage genome.The previous studies found that the expression of pstS of cyanophage S-SM1 and P-SSM2 was upregulated during infection to Synechococcus WH8102 and Prochlorococcus NATL2A,respectively,under P-limited conditions.However,in the cyanophage genome,there is no gene encoding the transmembrane channel protein PstA/C.It revealed that PstS protein from cyanophage can interact with the transmembrane channel from the host,and form a heterologous ABC transporter.Here,we show the X-ray structure of the liganded form of the phosphate-binding protein(PstS)from cyanophage P-SSM2 and Syn19.In addition,we simulated a model of PstS-PstCA complex,a heterologous ABC transporter,through which cyanophage PstS binds to the transmembrane channel PstCA from the host to form a chimeric transporter for the uptake of phosphate.The key residues involved in interface formation are highly conserved,and the cyanophages could be classified via the specific residues towards the corresponding hosts.This study gave us some information for the co-evolution between cyanobacteria and their cyanophages,and provided some hints for controlling the harmful algal bloom due to water eutrophication.