Crystallographic Study Of The Transmembrane Transport Channel Protein BamA Of Borrelia Burgdorferi

The cell membrane of Gram-negative bacteria are composed of inner and outer membrane structures.Transmembrane channel proteins in the outer membrane play an extremely important role in the material exchange between the bacteria and the external environment.Theβ-barrel assembly machinery（BAM）is one of the important outer membrane channel protein complexes in which theβ-barrel assembly machine A（Bam A）is its core channel protein.Mainly composed of a C-terminal beta-barrel domain and an N-terminal POTRA domain containing one or more polypeptide transport-related domains,its homologous proteins exist not only in the outer membrane of prokaryotes,but also in the outer membrane of organelles of mitochondria and chloroplasts of eukaryotes.BorreliaBurgdorferi,Bb is a pathogenic spirochete of Lyme disease,and its BamA（BbBamA）is different from prokaryotic Gram-negative bacteria Bam A and eukaryotic chloroplast BamA（homology is less than 20%）in amino acid primary sequence.It is speculated that the protein should have a unique three-dimensional spatial structure.However,at present,the precise three-dimensional structure of BbBamA is not clear,and the evolutionary relationship between the three BamA proteins in structure is not clear.In this study,three POTRA domains（BbBamA-POTRA P1-P3）at the N-terminal of BbBamA were selected as the research objects,and BbBamA-POTRA P1-P3 was successfully cloned,expressed,purified,crystallized and preliminary X-ray diffraction analysis,which laid a foundation for further structural analysis and functional analysis.The main findings are as follows:1.Under the condition of heterologous expression,BbBamA-POTRA P1-P3recombinant protein which can be used for protein crystallization was successfully obtained.2.High quality Bb BamA-POTRA P1-P3 protein crystals were successfully screened and optimized.3.High quality BbBamA-POTRA P1-P3 protein parent crystal data were successfully collected.Its belonged to the space group P2₁ with the highest resolution of 2.00?.The Matthews coeffificient was calculated to be 2.92?³Da^-1,assuming the presence of two molecules per asymmetric unit,and the corresponding solvent content was 57.9%.