Protein Purity
≥85% by SDS PAGE
Extinction Coeff
A280 nm = 10.16 at 1.0 mg/ml for pure C3
Molecular Weight
~185,000 Da (2 chains)
General Description
Rabbit C3 is purified from pooled normal rabbit serum. C3 is central to the activation of all three pathways of complement activation (Law, S.K.A. and Reid, K.B.M. (1995)). Initiation
of each pathway generates proteolytic enzyme complexes (C3 convertases) which are bound to the target surface. These enzymes cleave a peptide bond in C3 releasing the anaphylatoxin C3a and activating C3b. For a brief time (~60 µs) this nascent C3b is capable of reacting with and covalently coupling to hydroxyl groups on the target surface. Carbohydrates are the favored target, but protein hydroxyls and amino groups also react. This process of tagging the target surface with C3b is called opsonization. The reactive site in nascent C3b is a thioester (Tack B.J., et al. (1980); Pangburn M.K. and MüllerEberhard H.J. (1980)) and C3b is linked to the target through a covalent ester bond (an amide bond is formed if C3b is attached to amino groups). Most of the C3 activated during complement activation never attaches to the surface because its thioester reacts with water forming fluid phase C3b which is rapidly inactivated by factors H and I forming iC3b. Surface-bound C3b is necessary in all three pathways for efficient activation of C5 and formation of C5b-9 complexes that lyse the target cell membrane. Surface-bound C3b and its breakdown products iC3b and C3d are recognized by numerous receptors on lymphoid and phagocytic cells which use the C3b ligand to stimulate antigen presentation to cells of the adaptive immune system. The end result is an expansion of target-specific B-cell and T-cell populations.
Physical Characteristics & Structure
Analysis of purified rabbit C3 by SDS/polyacrylamide gel electrophoresis under non-reduced conditions shows its mobility to be similar to that of human C3 (185,000 daltons). Under reduced conditions, migration of the rabbit C3 alpha chain is similar to that of human (110,000 daltons) while the rabbit beta chain migrates slightly ahead of the human (75,000 daltons) but behind that of mouse (62,000 daltons). The extinction coefficient (E1%/280nm) for rabbit C3 was not calculated as the amino acid sequence only for the alpha but not the beta chain is available. To determine the concentration of purified rabbit C3 the extinction coefficient (E1%/280nm) was assumed to be 10.0.
Function
The biological functions of C3 are described above in the General Description section.
Genetics
The NCBI Gene ID number for rabbit C3 is 100009121 and the UniProt accession number for rabbit C3 alpha chain is P12247.
Precautions/Toxicity/Hazards
This protein is purified from animal plasma/serum and therefore precautions appropriate for handling any animal blood-derived product must be used.
References
Law, S.K.A. and Reid, K.B.M. (1995) Complement 2nd Edition (ISBN 0199633568) Oxford University Press, Oxford.
Tack BF, Harrison RA, Janatova J, Thomas ML, Prahl JW. (1980) Evidence for presence of an internal thiolester bond in third component of human complement. Proc Natl Acad Sci U S A. 77:5764-8.
Pangburn M.K. and Müller-Eberhard H.J. (1980) Relation of putative thioester bond in C3 to activation of the alternative pathway and the binding of C3b to biological targets of complement. J Exp Med. 152:1102-14.
