Characterization of cofactor activity involves the protein interacting with C3b or C4b, molecules that can be cleaved by factor I (FI) to produce the inactive forms iC3b, C3c, C3dg, C4d and C4c. Therefore, incubating the cofactor and FI with the substrates C3b or C4b and assessing protein cleavage by SDS-PAGE is a simple and common strategy to demonstrate cofactor activity. This method plays a crucial role in the characterization of fluid-phase cofactors, such as factor H (FH) and C4bp, or cell-associated cofactors, such as membrane cofactor protein (MCP) and complement receptor 1 (CR1). Creative Biolabs has long been committed to complement research. Equipped with world-leading technology platforms and professional scientific researchers, our expert team can assist you in designing the best customized research plan to meet the requirements of your complement project.
Cofactor Activity Assay Protocol
Here, we are willing to share a simple protocol for cofactor activity assay.
Fig.1 Flow chart cofactor activity assay.
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Published Data
Fig. 2 Structure of CD46.1
CD46 provides protection to autologous cells against complement-mediated damage by binding to and functioning as a significant cofactor for complement proteins C3b and C4b. Studies have shown that CD46 also plays a role in mediating adaptive immune responses and promoting autophagy. In addition to these physiological roles, numerous pathogens—including certain adenoviruses, human poxvirus 6 (HHV-6), streptococci, and Neisseria—employ CD46 as a receptor for cellular entry. To enhance understanding of the interactions between CD46 and its diverse ligands, the three-dimensional structure of an extracellular fragment of CD46 comprising all four short consensus repeat (SCR) domains (CD46-4D) was elucidated. The structure provides the basis for identifying the binding sites of several CD46 ligands that bind to the C-terminal region of the protein. It also reveals an unexpected kink between domains SCR3 and SCR4 that affects the conformation of CD46 on the cell surface and the recognition of its ligands.
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Reference
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Persson, B. David, et al. "Structure of the extracellular portion of CD46 provides insights into its interactions with complement proteins and pathogens." PLoS Pathogens 6.9 (2010): e1001122. Distributed under Open Access license CC BY 4.0, without modification.
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