Classical complement pathway component C1q: purification of human C1q, isolation of C1q collagen-like and globular head fragments and production of recombinant C1q-derivatives. Functional characterization.
The classical complement pathway (CCP) activation is a multimolecular complex, composed of three subcomponents namely C1q, C1r, and C1s. C1q is the recognition subunit of this complex and its binding to the specific targets leads to the formation of active C1, which in turn activates the CCP in an immunoglobulin-dependent or -independent manner. C1q is a hexameric glycoprotein composed of 18 polypeptide chains of three different types (A, B, and C), organized in two fragments-collagen-like (CLR) and globular head (gC1q) possessing different functional activity. The contemporary knowledge of the C1q structure allows the isolation and purification of a C1q molecule from serum by combination of different chromatography procedures including ion-exchange, size-exclusion, and affinity chromatography, as well as the isolation of CLR and gC1q by limited enzymatic hydrolysis of the native C1q molecule. In this chapter, we described methods for purification of human C1q and its CLR and gC1q fragments, as well as methods for their biochemical and functional characterization. The production and purification of recombinant C1q derivatives ghA, ghB, and ghC (globular fragments of the individual C1q chains) are also presented.
Department of Biochemistry, Sofia University "St. Kliment Ohridski", Sofia, Bulgaria.
Mihaela
M
Kojouharova
D002845:Chromatography / Q000379:methods; D002851:Chromatography, High Pressure Liquid; D015922:Complement C1q / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D003171:Complement Pathway, Classical; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006461:Hemolysis / Q000276:immunology; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D011485:Protein Binding / Q000276:immunology
D007074:Immunoglobulin G; D010446:Peptide Fragments; D015922:Complement C1q