title: Heterologous expression and purification of biologically active domains 3 and 4 of human polymeric immunoglobulin receptor and its interaction with choline binding protein A of Streptococcus pneumoniae.
abstract: Streptococcus pneumoniae, one of the common causes of pneumonia, colonises the epithelium via the interaction between a choline binding protein of S. pneumoniae and the human polymeric immunoglobulin receptor (pIgR). One of the functions of pIgR is to mediate the transcytosis of polymeric immunoglobulins from the basolateral to the apical surface of epithelial cells. S. pneumoniae invades human epithelial cells by exploiting the transcytosis machinery. Due to an increase in the prevalence of antibiotic resistant strains of S. pneumoniae, and the limitations and expense of the vaccines available, extensive research may provide insights into the potential of new therapeutic regimes. This study investigated the potential of pIgR domains as an alternative non-antibiotic immune therapy for treating pneumonia. The aim was to determine the binding affinity of recombinant D3D4 protein, the domains of pIgR responsible for binding S. pneumoniae, to recombinant R1R2 repeat domains of choline binding protein A of S. pneumoniae. Biologically active recombinant D3D4 was produced in Escherichia coli using a gel filtration chromatography refolding method, a novel approach for the refolding of pIgR domains, after the purification of inclusion bodies using nickel affinity chromatography. Surface Plasmon resonance (SPR) spectroscopy showed that purified recombinant D3D4 binds recombinant R1R2 with an equilibrium dissociation constant (KD) of 3.36×10(-7)M.
authors: Department of Biochemistry and Microbiology, Nelson Mandela Metropolitan University, P.O. Box 77000, Port Elizabeth 6031, South Africa.
authors: Luanne
authors: L
authors: Venables
authors: Sharlene
authors: S
authors: Govender
authors: Vaughan
authors: V
authors: Oosthuizen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D058849:Protein Refolding; D017434:Protein Structure, Tertiary; D019056:Receptors, Polymeric Immunoglobulin / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D016415:Sequence Alignment; D020349:Surface Plasmon Resonance
chemical_list: D001426:Bacterial Proteins; D019056:Receptors, Polymeric Immunoglobulin; D011994:Recombinant Proteins; C110344:SpsA protein, Streptococcus pneumoniae
keywords: Choline binding protein A; Polymeric immunoglobulin receptor; Streptococcus pneumoniae; Surface plasmon resonance spectroscopy