Complement cofactor

Every regulatory system in the body must be prevented from overactivity or activity that is unnecessarily prolonged. This can help us understand that, just as with blood clotting (Fig. 12-17), a network of regulatory factors controls the complement system. Among these are an inhibitory C4b-binding protein (C4BP),181 which acts to prevent excessive formation of the C4b C2a complex (Fig. 31-8). Complement cofactor I is a serine protease that cleaves both C3b and C4b into smaller pieces in the presence of cofactor... 

Dihydroflavonol 4-reductase (DFR EC 1.1.1.219) is a member of the short-chain dehydrogenase/reductase family and catalyzes the stereospecific conversion of (+)-(2R,3R)-dihydroflavonols to the corresponding (2R,3S,4S) flavan-3,4-cw-diols (leucoanthocyanidins), with NADPH as a required cofactor. The enzyme activity was first identified in cell suspension cultures of Douglas fir (Pseudotsuga menziesii) and was shown to be related to the accumulation of flavan-3-ols and proanthocyanidins [96]. Leucoanthocyanidins and DFR were later shown to be required for anthocyanidin formation by complementation of Matthiola incana mutants blocked between dihydroflavonol and anthocyanidin biosynthesis [97, 98], DFR has been purified to apparent homogeneity and biochemically analyzed from flower buds of Dahlia variabilis [99]. DFR was shown to accept different substrates depending on the plant species from which it was isolated (reviewed in 100). 

Hepatocytes Culture methods available Full enzyme complement - contain enzymes and enzyme cofactors not present in subcellular fractions Full enzyme complement - contain enzymes and enzyme cofactors not present in subcellular fractions Can be used quantitatively Useful for prediction of Phase 1 and 11 metabolism Require fresh tissue No cell-cell contact Need collagenase digestion Handling difficult Closed system so may not be representative of in vivo situation... 

Figure 2. Immune lysis of a sensitized liposome. Immobilized antibody-sensitized liposome undergoes complement-induced lysis. Released enzyme catalyzes substrate-product reaction with concomitant reduction of immobilized cofactor. The cofactor is electrochemi-cally reoxidized and the current is related to the analyte concentration. (See text for discussion.) Symbols A, analyte (antigen) Y, antibody S, sub-...

Figure 7-2 Control of the alternative complement pathway by activating surfaces. When complement component C3b binds to a surface, there exist two possible outcomes. Under normal conditions, when no activating surface is present (e g., if C3b has contacted normal tissue), sequential addition of blood cofactors H and I converts C3b into C3c, inactivating the complement protein. If an activating surface such as a microbe or damaged tissue is encountered, sequential addition of factors B and D drives the alternative pathway to the normal properdin (P) intermediate, and the complement cascade is triggered. The properdin-containing component (C3bBbP) feeds back to the beginning of the pathway, generating more C3.

P (properdin) can also bind to the C3 convertase. Its role is to stabilize the complex and hence it is considered a cofactor-activator in the alternative pathway. These components plus additional C3b molecules form the C5 convertase, an enzyme complex that peoteolytically converts C5 to C5a and CSb. Properdin stabilizes C3b and Bb in the complex and protects these proteins from proteolytic inactivation by factor I. Factor H competes for Bb in the C5 convertases, the same as it does in the C3 convertase. The alternative pathway has also been called the properdin pathway because of properdin s participation in alternative pathway C3 and C5 convertases. C5b is a component in the terminal complex of the complement activation process, the MAC. The MAC is composed of a self-assembled, noncovalent complex of C5b, C6, C7, C8, and C9. Together these components produce a pore-like structure that makes the membrane of the cell to which it is attached permeable and causes cell death. Under the electron microscope the MAC appears like an impact crater similar to those observed on the surface of the moon. C5a is also an anaphylatoxin like C3a, but it is more potent. C5a is also a chemokine and attracts phagocytic cells to the site of complement activation. 

Because inactivation of factors Va and Villa by activated protein C promotes the dissociation of the pro-teinases, cofactor protein inactivation complements the action of the proteinase inhibitors. This eliminates the protection that the proteinases have when bound to their cofactor proteins and substrates. Inactivation of proteinases by SERPINS occurs via a common mechanism that involves a Michaelis complex between the proteinase and the inhibitor (Figure 36-16). This mechanism applies to all serine proteinases of the hemostatic system, i.e., the procoagulant, anticoagulant, and fibrinolytic subsystem proteinases. 

The PS-I core complex (CC 1) used by these workers for crystallization and X-ray crystallographic analysis was purified from Synechococcus elongatus and found to exist in the trimeric form. Each PS-1 monomer unit consisted of the major PsaA and PsaB polypeptides, plus several other smaller subunits, and included a full complement of electron-transfer cofactors, P700, (A), Ao, A, FeS-X and FeS-A/B. It was also known to contain 90 Chl-a molecules per P700. The amount of core-antenna chlorophyll-a in this preparation was comparable to that found in the core complexes of spinach and other cyanobacteria. To simplify the designation, the core-antenna Chi a will now be abbreviated as CA-Chl a. ... 

Effectors accelerate (activators) or block (inhibitors) the catalytic process. They are bound to the enzyme rather loosely and may therefore easily dissociate. Many of them are metal ions, e.g. Mg++) Ca++, Zn++, K+) and Na+, which either form stoichiometric complexes with the substrate, stabilize an optimal protein conformation, or effect the association of subunits. These inorganic complements of enzyme reactions are frequently subsumed together with coenzymes as cofactors. 

Mathews, I. I., A. M. Deacon, J. M. Canaves, D. McMullan, S. A. Lesley, S. Agar-walla, and P. Kuhn. Functional analysis of substrate and cofactor complex structures of a thymidylate synthase-complementing protein. Structure 11 677-90. 

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