Expression,Purification And Crystallization Of Unconventional HLA-B27 Homodimer

Ankylosing spondylitis is a sacroiliac joint and spine attachment inflammation as the main symptoms of the disease.It is a large part of the limbs,as well as intervertebral disc annulus and its adjacent connective tissue fibrosis and ossification,and joint stiffness as the characteristics of chronic inflammatory disease.Ankylosing spondylitis is belong to rheumatism and seronegative spondyloarthropathy.The cause of the disease is not clear.HLA antigen is a product of the major histocompatibility complex(MHC)expression.It is mainly responsible for the recognition among cells,inducing immune response and regulating the immune response functionin the immune system.According to the structure and function of HLA antigen and the distribution of tissue,it was divided into three categories.Class I molecules are HLA-A,-B and-C series antigens,which are widely distributed on the surface of nucleated cells.HLA-B27 is a human leukocyte antigen,belonging to one of the HLA-B loci.In the spondyloarthropathy,ankylosing spondylitis(AS)is highly correlated with HLA-b27,but the mechanism of treatment is not yet clear.The theory of HLA-b27 homologous dimerization induced-desease has recently become more and more popular,but lack of structural data which could be analyzed and confirmed.The structure of HLA-B27 homodimer is expected to elucidate the molecular mechanism of ankylosing spondylitis from the structural point of view and provide a theoretical basis for subsequent drug design and allergy prevention.In this study,we try to obtain the protein with high purity and homogeneity through prokaryotic expression,in vitro refolding and purification,and try to obtain crystals which can be used for X-ray diffraction by crystal selection.In this study,E.coli expressed HLA-B27 protein as inclusion body.Through molecular sieve and SDS-PAGE analysis,it was found that the protein was a large amount of polyol and a small amount of dimer after renaturation.We finally obtained 0.2 mg of HLA-B27 homodimer with high purity and homogeneity from 20 mg HLA-B27 inclusion body,and observed the crystal in the primary crystal screening,which laid an important foundation for obtaining protein crystals and structural data.and screened protein crystals in primary screening.