As shown in the alternative pathway uses three different recognition molecules to identify the host or targets. Factor H is a protein composed of twenty small domains arranged like beads on a flexible string (). The N-terminal four domains regulate the complement amplification system [25
]. The other sixteen domains contain a variety of binding sites which control the functional effectiveness of domains 1–4 at the N-terminus. At least two sites interact with host cell surface markers, thought to be polyanions, and multiple C3b-binding sites interact with cell-surface deposited C3b/C3d [26
], allowing factor H to effectively control the spontaneous activation of the alternative pathway on host surfaces. Complement-mediated pathology in the host can be triggered by the lack of proper recognition of markers on host cells due to mutations in the binding sites on factor H, as evidenced by inherited atypical hemolytic uremic syndrome (aHUS) [34
] and age-related macular degeneration (ARMD) [35
]. The specific chemical nature of host markers has yet to be reported, but both regions (domain 7 and 19–20, ) are thought to interact with host polyanionic molecules, such as sialic acids and glycosoaminoglycans. Complement activating organisms, e.g.
yeast, coated with heparin cease to activate human complement [36
]. On the other hand, sheep cells, which do not activate human complement, activate the alternative pathway after surface sialic acid is removed [37
]. Thus, although the human counterparts of these polyanions have never been identified they are assumed to be polyanionic in nature. The primary difficulty in identification appears to be the low affinity between individual markers and factor H. In vivo
factor H may utilize both surface-bound C3b and multiple host specific contacts to identify host [39
]. On surfaces lacking markers recognized by factor H (bacteria, fungi, agarose, glomerular basement membrane, etc.) alternative pathway-mediated C3b amplification proceeds unabated due to the ineffectiveness of factor H on such surfaces.
Figure 2 Host/target discriminatory sites on factor H. While the N-terminal four domains express all of the complement regulatory functions of factor H, this activity is controlled by other polyanion and C3b binding sites localized to domains 7, 12–13 (more ...)
Properdin is one of the six proteins involved in activation of the alternative pathway. It stabilizes the central enzyme in the amplification process and thus accelerates activation [41
]. Although first identified in 1959 [42
], it was not until 2007 [17
] that it was shown to be a pattern recognition molecule capable of binding to Neisseria gonorreae
, necrotic human cells, and fungi and initiating activation of the alternative pathway [16
]. People with properdin deficiencies have recurrent episodes of meningococcal [43
] infections suggesting that recognition and elimination of these organisms are uniquely dependent on properdin. Certain molecular structures recognized by properdin have been proposed, such as cell surface GAGs, DNA, as well as non-sulfated glycoconjugates [16
C3b attaches covalently to targets being attacked by all three pathways of complement as illustrated in . This critical step in complement activation is not normally thought of as a target recognition event, however, C3b exhibits strong preferences () among different sugars (carbohydrates are its preferred attachment site on most targets) and an even stronger preference for amino acid hydroxyl groups (Thr, Ser and Tyr) when they are available [47
]. Attachment to xylose is preferred 12-fold over attachment to inositol and two fold over attachment to glucose (). Even preferences for particular hydroxyl positions on sugars have been demonstrated [49
]. Attachment to the phenyl hydroxyl group of tyrosine is preferred 50-fold over glucose and the ester link to amino acids is more stable than that to sugars. In one case it was shown () that a series of yeast mutants capable of synthesizing surface polysaccharides terminating with one, two, three or four xylose sugars (the most highly preferred sugar for C3b attachment) exhibited a 4-fold difference in C3b attachment efficiency [49
]. The rate and extent of complement activation by these yeast also reflected this quantitative difference suggesting that the specificity measured with sugars is relevant to complement activation [49
Specificity of C3b Attachment
Figure 3 The specificity of the reactive thioester site of C3 for carbohydrates can determine the rate and extent of complement activation. Different strains of yeast (Cryptococcus neoformans) capable of attaching different numbers of xylose residues per unit (more ...)
Antigen-processing cells of the adaptive immune system utilize C3b/C3d-tagged antigens as their preferred antigens, through receptors CD21 and the BCR [52
], especially during the initial low antigen concentration phase of infections. This suggests that the chemical reactivity of C3b for particular sites on an antigen may be an important step governing antigen selection. It may also be important in regulating the pathway of cellular processing (Th1 versus
]. The recent discovery of a completely C3 deficient child with severely impaired B-cell, dendritic cell and T-cell responses supports this conclusion [55
]. The fact that the ability of activated C3 to attach to invaders occurred early in evolution suggests that the mechanism of invader identification and tagging by C3 might be an important factor in the selection of antigens. It seems reasonable to expect that the selection of specific structures on target surface molecules by C3b, and the covalent tag it attaches, will influence what polysaccharides and protein fragments are utilized for processing by antigen-presenting cells.
All three pathways of complement activation deposit C3b on their respective targets. If that surface lacks host markers or membrane-bound complement regulatory proteins (i.e., DAF, MCP, CR1, CD59) the alternative pathway will amplify the number of C3b on that surface. This system is capable of depositing millions of C3b protein molecules on a strong activator within five to 15 minutes of first contact with host blood. A single cell will elicit such a response suggesting that during early, low antigen dosage responses the molecules identified and tagged by C3b may have a disproportionate influence on what antigens get processed.