Structural And Functional Analysis Of NdhK In Synechocystis Sp. PCC6803

About20years ago, a novel cyanobacterial photosynthetic membrane proteincomplex, NADPH dehydrogenase (NDH-1), was discovered in the thylakoidmembrane. So far, the NDH-1complex consists of at least18subunits, i.e., NdhA toNdhS. ndhK is placed in an operon with ndhC and ndhJ. ndhK can transcribe rapidlywhen induced by high light and high CO2, so it must have a promoter that canresponse the change of the external environment to regulate gene expression.In order to understand the structure and function of ndhK promoter, ifrstly，weifnd out ndhK、s transcription initiation site is located at160bp upstream of the startcodon. Meanwhile, we found four possible promoters using bioinformatics analysis.After this, we construct a promoter probe vector which can be used in E.coli anddesign a series of promoter fragments for further analysis of promoter and regulatoryregions of ndhK. The analysis reveals that-374?-274bp，-438?-374bp，-604?-543bp，+1?+52bp in upstream of ndhK can promte gene expression and-543?-440bp can inhibite.Then，we transform some promoter fragments into wild Synechocystis sp. StrainPCC6803using pPT6803-l and get five mutant strains. The growth curve of thesemutant strains and WT are similar, and the bioluminescence levels from mutantstrains and WT are identical with the result get from the promoter probe vector, itproved that the results we get are right.In summary, we found:(l)the transcription initiation site of ndhK;(2)four possible promoter found by bioinformatics analysis;(3)four sequences which promote gene expression and one which inhibite;(4) constructing a promoter probe vector that can detect the activity of promoter fragment in E. coli.