Study On The Heterologous Expression Of LHI Protein Mutants In The Photorcactive Center Of Thermochromatium Tepidum

Thiophilic purple Thermochromatium(Tch.)tepidum is a photosynthetic bacterium.The light-harvesting pigment protein complex of Thermochromatium(Tch.)Tepidum is composed of a reaction center and a Light Harvesting Complex.Because of the specificity of light-harvesting LHI structure,its the spectral scanning absorption peak appears at 915 nm.This subject is mainly the heterologous expression of LHI protein mutants in photoreactive centers.Rhodospirillum(R.)rubrum belongs to purple non-sulfur photosynthetic bacteria.As the oxygen content decreases,an intracellular membrane(ICM)with a photosynthetic apparatus is synthesized.This apparatus is composed of a light-harvesting protein and a photoreaction center.Using molecular biology and spectroscopic analysis techniques,we cloned the Thermochromatium(Tch.)tepidum pufBA sequence in vitro and perform four mutations(alanine mutation,leucine mutation,tryptophan mutation and Aspartate mutation).The mutated pufBiAi sequence was constructed on the pPUTerm recombinant expression vector,and the expression plasmid was introduced into the mutated Rhodospirillum(R.)rubrum H2,the optimal inducing conditions of the mutant zygote were optimized,and the characteristics of the expressed protein were analyzed.The results showed that the mutated Thermochromatium(Tch.)tepidum pufBiAi could be induced to express normally in Rhodospirillum(R.)rubrum H2 and the expression level was stable.The results of UV-scanning showed that photosynthetic bacteria had characteristic absorption peaks near 890 nm,resulting in a blue-shift phenomenon of absorption peaks.The heterologous expression of Thermochromatium(Tch.)tepidum LHI pufBiAi protein facilitates the exploration of the crystal structure of Thermochromatium(Tch.)tepidum LHI,and provides a theoretical basis for further exploration of site-directed mutant proteins in vitro.The expression of the light-harvesting pigment protein heterologous membrane protein of Thermochromatium(Tch.)tepidum photosynthetic bacteria is helpful for the development of a novel membrane protein expression test system,and laid the foundation for the study of the crystal structure of the light-harvesting pigment protein.