Studies On The Heterologous Expression Of Light-Harvesting Complex I (LHI) From Thiorhodovibrio Sp.970

Photosynthetic bacteria Thiorhodovibrio sp.970 belongs to the Chromobacteriaceae of the photosynthetic bacteria.Its light-harvesting complex 1(LHI)has a maximum absorption peak near 970nm,which is currently the largest redshift among photosynthetic bacteria containing chlorophyll a,at present;In addition to the common pufBALMC gene sequence,the light-harvesting protein LHI of Thiorhodovibrio sp.970 also contains a second pufBA gene sequence downstream of pufC.The photosynthetic bacterium Rhodospirillum rubrum(R.rubrum)is a purple non-sulfurized facultative anaerobe.R.rubrumH2 is a mutant strain of R.rubrum,which cannot independently form cytoplasmic intracytoplasmic membrane(ICM),but can produce ICM under the stimulation of exogenous genes and encapsulate exogenous proteins to express in ICM.Therefore,in this study,two pufBA genes encoding LHI in Thiorhodovibrio sp.970 strain were heterologously expressed in R.rubrumH2 respectively,and the changes of its characteristic absorption peaks were observed.In this project,the heterologous expression of Thiorhodovibrio sp.970 light-harvesting complex LHI was studied.The pufB₁A₁ and pufB₂A₂ of Thiorhodovibrio sp.970 light harvesting complex LHI were cloned in vitro by molecular biology principle,the recombinant expression vectors of pPUCTerm-pufB₁A₁ and pPUCTerm-pufB₂A₂ were constructed by gene recombination technology,and the successfully constructed expression vectors were transformed into R.rubrumH2 through E.coli WM3064 binding transformation,PCR and SDS-PAGE were used to identify whether pufB₁A₁ and pufB₂A₂ genes were successfully expressed in R.rubrumH2,and the characteristic absorption peaks of the recombinant strains were detected by spectrophotometry.The results showed that the recombinant expression vectors pPUCTerm-pufB₁A₁ and pPUCTerm-pufB₂A₂were successfully constructed and expressed in R.rubrumH2,and the characteristic absorption peaks of the two recombinant strains were measured at 970 nm.These results indicate that pufB₁A₁ and pufB₂A₂ can be heterologous expressed separately in R.rubrumH2,and both have characteristic absorption peaks at 970 nm.The successful heterologous expression of the light-harvesting complex LHI in photosynthetic bacteria Thiorhodovibrio sp.970 is conducive to study the amino acid mutants of the light-harvesting complex LHI binding to Ca²⁺,and also lays a foundation for further research on the structure and spectral absorption of the light-harvesting complex LHI.