The lectin pathway of complement is activated when a carbohydrate recognition complex and associated serine proteases binds to the surface of a pathogen. Three recognition subcomponents have been shown to form active initiation complexes: mannan-binding lectin (MBL), L-ficolin, and H-ficolin. The importance of MBL in antimicrobial host defense is well recognized, but the role of the ficolins remains largely undefined. This report shows that L-ficolin specifically binds to lipoteichoic acid (LTA), a cell wall component found in all Gram-positive bacteria. Immobilized LTA from Staphylococcus aureus binds L-ficolin complexes from sera, and these complexes initiate lectin pathway-dependent C4 turnover. C4 activation correlates with serum L-ficolin concentration, but not with serum MBL levels. L-ficolin binding and corresponding levels of C4 turnover were observed on LTA purified from other clinically important bacteria, including Streptococcus pyogenes and Streptococcus agalactiae. None of the LTA preparations bound MBL, H-ficolin, or the classical pathway recognition molecule, C1q.

Mannan-binding lectin-associated serine protease 2 (MASP-2) has been described as the essential enzyme for the lectin pathway (LP) of complement activation. Since there is strong published evidence indicating that complement activation via the LP critically contributes to ischemia reperfusion (IR) injury, we assessed the effect of MASP-2 deficiency in an isogenic mouse model of renal transplantation. The experimental transplantation model used included nephrectomy of the remaining native kidney at d 5 post-transplantation. While wild-type (WT) kidneys grafted into WT recipients (n‫)7؍‬ developed acute renal failure (control group), WT grafts transplanted into MASP-2-deficient recipients (n‫)7؍‬ showed significantly better kidney function, less C3 deposition, and less IR injury. In the absence of donor or recipient complement C4 (n‫,)7؍‬ the WT to WT phenotype was preserved, indicating that the MASP-2-mediated damage was independent of C4 activation. This C4-bypass MASP-2 activity was confirmed in mice deficient for both MASP-2 and C4 (n‫,)7؍‬ where the protection from postoperative acute renal failure was no greater than in mice with MASP-2 deficiency alone. Our study highlights the role of LP activation in renal IR injury and indicates that injury occurs through MASP-2-dependent activation events independent of C4.

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Ficolins and mannan-binding lectins (MBLs) are the first components of the lectin branch of the complement system. They comprise N-terminal collagen-like domains and C-terminal pathogen-recognition domains (fibrinogen-like domains in ficolins and C-type carbohydrate-recognition domains in MBLs), which target surface-exposed N-acetyl groups or mannose-like sugars on microbial cell walls. Binding leads to activation of MBL-associated-serine protease-2 (MASP-2) to initiate complement activation and pathogen neutralisation. Recent studies have shown that MASP-2 binds to a short segment of the collagen-like domain of MBL. However, the interaction between ficolins and MASP-2 is relatively poorly understood. In this study, we show that the MASP-2 binding site on rat ficolin-A is also located within the collagen-like domain and encompasses a conserved motif that is present in both MBLs and ficolins. Characterisation of this motif using site-directed mutagenesis reveals that a lysine residue in the X position of the Gly-X-Y collagen repeat, (Lys56 in ficolin-A) which is present in all ficolins and MBLs known to activate complement, is essential for MASP-2 binding. Adjacent residues also make important contributions to binding as well as to MASP activation probably by stabilizing the local collagen helix. Equivalent binding sites and comparable activation kinetics of MASP-2 suggest that complement activation by ficolins and MBLs proceeds by analogous mechanisms.

Mannan-binding lectin (MBL) initiates complement activation by binding to arrays of carbohydrates on the surfaces of pathogenic microorganisms and activating MBL-associated serine proteases (MASPs). Separate point mutations to the collagenous domain of human MBL are associated with immunodeficiency, caused by reduced complement activation by the variant MBLs as well as by lower serum MBL concentrations. In the work reported here, we have used the well characterized rat lectin pathway to analyze the molecular and functional defects associated with two of the variant proteins. Mutations Gly25 → Asp and Gly28 → Glu create comparable structural changes in rat MBL but the G28E variant activates complement >10-fold less efficiently than the G25D variant, which in turn has ∼7-fold lower activity than wild-type MBL. Analysis of mutant MBL · MASP-2 complexes assembled from recombinant components shows that reduced complement activation by both mutant MBLs is caused by failure to activate MASP-2 efficiently on binding to a mannan-coated surface. Disruption of MBL-MASP-2 interactions as well as to changes in oligomeric structure and reduced binding to carbohydrate ligands compared with wild-type MBL probably account for the intermediate phenotype of the G25D variant. However, carbohydrate binding and MASP-2 activation are ostensibly completely decoupled in complexes assembled from the G28E mutant, such that the rate of MASP-2 activation is no greater than the basal rate of zymogen MASP-2 autoactivation. Analogous molecular defects in human MBL probably combine to create the mutant phenotypes of immunodeficient individuals.