3. Compl e ment What is “it”? A series of serum proteins. How is it recognized? By its ability to mediate cell lysis. (Review the fascinating story of its discovery!) What does it have to do with IMMUNOLOGY?

4. The overview…. Complement has three functions: –Opsonin –Chemoattractant –Membrane Attack Complex (MAC) Complement functions in two (three?) systems: –Alternative –Classical –Lectin-based

5. The alternative system C3 is the principal protein of concern. –Exists in high concentrations (1.2 mg ml -1 –Contains an unstable internal thiolester with slow “spontaneous” (or serum protease mediated) cleavage. By convention, the smaller product is designated “a” and the larger “b” (though there is one exception.) C3b can bind to bacteria and enhance phagocytosis.

7. The alternative system The production of C3b can be catalyzed by “C3 convertase.” C3 convertase is produced by C3b associating with factor B in the presence of Mg ++ to form C3bB which in the presence of factor D is cleaved to C3[bBb]. Nota bene: C3[bBb] is C3 convertase. {Here brackets substitute for a horizontal bar above the letters bBb; by convention, the bar over the letters means that the complex has enzymatic activity.}

8. The alternative system The production of C3b can be catalyzed by “C3 convertase.” Because C3 convertase both creates and contains C3b, the complex mediates a “positive feedback” system. Thus, the reaction has the potential “to run away” or “explode.” Factor H can displace factor B. The C3bH complex is the substrate of factor I which inactivates C3b by cleaving it into C3d and C3g. C3[bBb] is C3 convertase.

9. The alternative system C3b has FOUR fates: (i) it can bind to a microbial surface; (ii) it can associate with factor B; (iii) it can associate with factor H; or (iv) it can bind another C3b. (What happens here?) The half-life of C3b is approximately 100 microseconds ( so the “choices” occur quickly!).

10. The alternative system If C3[bBb] binds a second C3b, it becomes C3[bBb]3b which is “C5 convertase”. C5 convertase cleaves C5 into C5a and C5b. C5b is unstable and is stabilized by C6. C5b6 allows C6 and C7 to associate and penetrate a membrane. The C5b67 recruits C8 which organizes C9 to form a “membrane attack complex” (MAC).

11. The alternative system C5 convertase cleaves C5 into C5a and C5b. C5b is unstable and is stabilized by C6. C5b6 allows C6 and C7 to associate and penetrate a membrane. The C5b67 recruits C8 which organizes C9 to form a “membrane attack complex” (MAC).

12. The alternative system Finally…. –C–C3a and C5a are –C–CHEMOATTRACTANTS! Think of the multiple dimensions of this system which, incredibly, is the product of evolution. A great puzzle is how such interdependent systems can emerge incrementally.

13. The CLASSICAL pathway Stimulated by antibodies: specifically: IgM and IgG (subclasses 1, 2, 3) Start with C1q a HUGE protein (410,000 daltons!) Composed of 18 peptides. Peptides can associate to form trimers; six sets of trimers make C1q. C1q has helical “stalks” and globular “heads.” (N. B. the heads are the carboxy end and the stalks are the amino ends)

14. The CLASSICAL pathway Also associated with C1 are C1r and C1s which associate to make dimeric pairs (C1r 2 s 2 ); the dimeric pair joins C1q to form C1qr 2 s 2.

15. The CLASSICAL pathway C1qr 2 s 2 binds to TWO immunoglobulins. –The complement binding sites of circulating IgM are too far apart to bind complement; – only when IgM is bound does it fold so that C1qr 2 s 2 can “see” nearby complement binding domains. – IgG concentrations must be high in the vicinity of antigens for threshold levels of complement binding domains to be present.

16. The CLASSICAL pathway When C1qr 2 s 2 is bound to requisite number of immunoglobulins, C1r “autocatalytically” converts to C1[r]. C1[r], in turn, converts C1s to C1[s]. C1[s] cleaves C2 and C4. C4 is cleaved to C4a and C4b; C4b associates with its “target” which is C2. C2 is cleaved by C1[s] making C[4b2a] which is a C3 convertase! (Note that 2a is bigger than 2b, this nomenclature being the one exception to the convention that “a” is smaller than “b.”) As with the other C3 convertase, C3b can join C[4b2a] to make C[4b2a]3b which is also a C5 convertase.

17. The CLASSICAL pathway

20. The complement pathways…

21. The LECTIN pathway Lectins are proteins which bind to carbohydrates. Many bacteria have many mannose residues on their surface. The lectin-based complement system begins with a “mannose-binding protein” (MBP). MBP reacts, in turn, with a MBP-associated serine protease (MASP). MASP functions, in effect, like activated C1q[r 2 s 2 ], that is a C3 convertase.

22. The most amazing circumstance Erythrocytes (!) deliver the complex of antigen – antibody – complement to the liver and spleen for consumption by phagocytes.