Native C3 Research Articles

The effects of eastern red cedar (Juniperus virginiana) invasion and removal on a dry bluff prairie ecosystem

Eastern red cedar (Juniperus virginiana) establishment increased dramatically in the tallgrass prairie biome of North America during the last 30 years. Since many of the remaining remnant prairies occur on steep, dry, and nutrient poor sites, threatened by the invasion of native and non-native woody species, it is important to understand how an invasive species such as eastern red cedar influences key environmental factors that may determine the future trajectory of these systems and whether abiotic and biotic components of the system are resilient following cedar removal. To address these issues we: (1) investigated the influence of eastern red cedar on micro-environmental factors; (2) evaluated how these micro-environmental factors responded to eastern red cedar removal; and (3) assessed the effect of eastern red cedar on herbaceous species germination and distribution. The invasion of eastern red cedar was associated with lower surface light availability and soil temperature, as seen in prior studies, but otherwise had effects distinct from those observed in prior studies. There was no effect of cedar on soil pH, and unlike prior studies, cedar patches had higher soil moisture compared to native C4 prairie grass plots. Moreover, these effects had strong spatial signatures, with impacts of invasion on micro-environment and native vegetation differing dramatically with slope position and aspect. Three years after eastern red cedar was removed, micro-environmental factors and species composition became similar to the tree-free grass-dominated plots, indicating a significant capacity for recovery following possible cedar control. In a broader context, this study sheds light on the pathways and mechanisms driving the impacts of this biological invasion on dry, steep, nutrient poor systems and illustrates the capability of these systems to recover once the invading species is removed.

Melilotus officinalis (yellow sweetclover) causes large changes in community and ecosystem processes in both the presence and absence of a cover crop

Non-native species are hypothesized to decrease native species establishment and cover crops are hypothesized to decrease non-native species abundance. Although many studies have compared invaded to non-invaded habitats, relatively few studies have experimentally added non-native species to directly examine their effects. In a greenhouse mesocosm experiment, we tested the effects of non-native forbs (Melilotus officinalis, Verbascum thapsus, and Lespedeza cuneata), a proposed C3 grass cover crop (Pascopyrum smithii), and a commonly seeded non-native C3 grass (Bromus inermis) on the establishment of target native C4 prairie grass species. All treatments contained the same seed density of target C4 species and were begun on bare soil collected from the field. The legume M. officinalis strongly decreased the abundance of all other species, species diversity, and light and soil moisture levels. Surprisingly, M. officinalis took up relatively large amounts of labeled nitrogen (15N) from the soil early in its development, but M. officinalis fixed nitrogen, thus increasing nitrogen in biomass nearly fivefold by the end of the study. We found few effects of either C3 grass species on non-native forbs or C4 target species, but seeded P. smithii did increase species diversity. Non-native plants therefore impeded native C4 grass establishment through long-lasting effects of target species seedbank depletion (death of most target seedlings) and altered nutrient availability. The effects of M. officinalis were not reduced by the presence of a cover crop.

Persistence of Native C4 Grasses under High-Intensity, Short-Duration Summer Bison Grazing in the Eastern Tallgrass Prairie

Disturbances such as burning or grazing maintain the herbaceous nature of eastern tallgrass prairie. These disturbances are also known to affect the relative abundance of warm-season (C4) and cool-season (C3) grasses in native prairie. Although burning is a commonly used tool, the utility of livestock grazing to manage restored prairie is less understood. We established five monocultures and one mixture of C4 grass species of the eastern tallgrass prairie in southern Wisconsin. To examine their persistence under high-intensity, short-duration summer grazing, we estimated cover of several functional groups and C4 species over a 6-year period (2000 through 2006) in a randomized complete block design. After a 2-year establishment phase (1998-1999), bison were rotated through paddocks two or three times annually during late June, July, or early August. All C4 grasses declined over time but at different rates depending on the species. Switchgrass (Panicum virgatum) decreased at the lowest rate, whereas Little bluestem (Schizachyrium scoparium) cover declined faster than Big bluestem (Andropogon gerardii), Indiangrass (Sorghastrum nutans), and Sideoats grama (Bouteloua curtipendula), whose rates of decline were not significantly different from each other. Succession followed a predictable trajectory with annual grasses initially colonizing interstitial space among C4 grasses, followed by legumes, which ultimately gave way to exotic C3 forage grasses. The focal C4 grasses remained the dominant functional group 8 years postseeding, but recolonization by non-native C3 grasses increased over the study period.

Enzyme-independent, orientation-selective conjugation of whole human complement C3 to protein surfaces

Complement C3 is a central component of the humoral immune system. Upon triggering of the complement cascade, proteolytic fragments of C3 mediate important processes such as opsonization and lymphocyte activation. C3 possesses an internal thioester that mediates covalent attachment of proteolytically activated C3 to target surfaces. Treatment of native C3 with methylamine cleaves the thioester bond and exposes a free sulfhydryl group at the target-binding face of the protein. Through the use of sulfhydryl-reactive heterobifunctional cross-linking and biotinylation reagents, we demonstrate the capacity to form stable, multimeric whole human C3-protein conjugates in a fashion reflecting the orientation of physiologically-activated C3. We speculate that this C3 conjugation strategy presents a route for targeting dendritic cells and macrophages. In addition, manipulation of the thioester bond could enhance the study of biological roles of C3 and related proteins such as C4, and also of transmissible agents that exploit complement function such as prions.

Activation of Complement Component C5: COMPARISON OF C5 CONVERTASES OF THE LECTIN PATHWAY AND THE CLASSICAL PATHWAY OF COMPLEMENT

Although the initiating complex of lectin pathway (called M1 in this study) generates C3/C5 convertases similar to those assembled by the initiating complex (C1) of the classical pathway, activation of complement component C5 via the lectin pathway has not been examined. In the present study kinetic analysis of lectin pathway C3/C5 convertases assembled on two surfaces (zymosan and sheep erythrocytes coated with mannan (E(Man))) revealed that the convertases (ZymM1,C4b,C2a and E(Man)M1,C4b,C2a) exhibited a similar but weak affinity for the substrate, C5 indicated by a high K(m) (2.73-6.88 microm). Very high affinity C5 convertases were generated when the low affinity C3/C5 convertases were allowed to deposit C3b by cleaving native C3. These C3b-containing convertases exhibited K(m) (0.0086-0.0075 microm) well below the normal concentration of C5 in blood (0.37 microm). Although kinetic parameters, K(m) and k(cat), of the lectin pathway C3/C5 convertases were similar to those reported for classical pathway C3/C5 convertases, studies on the ability of C4b to bind C2 indicated that every C4b deposited on zymosan or E(Man) was capable of forming a convertase. These findings differ from those reported for the classical pathway C3/C5 convertase, where only one of four C4b molecules deposited formed a convertase. The potential for four times more amplification via the lectin pathway than the classical pathway in the generation of C3/C5 convertases and production of pro-inflammatory products, such as C3a, C4a, and C5a, implies that activation of complement via the lectin pathway might be a more prominent contributor to the pathology of inflammatory reactions.

Leaf morphology and anatomy in two contrasting environments for C3and C4grasses of different invasion potential

Leaf morphology, coarse structure and anatomy were compared for two invasive C4, two non-invasive C4, and two expanding native C3 grass species grown in their original, high-light semiarid temperate habitat, and in a growth room under variable moderate light and favourable supply of water and nutrients. It was hypothesised that (H1) among C4 grasses leaf structural response will be greater for invasive than for non-invasive species, and (H2) for plants of high spreading capacity C4 species will be less responsive than C3 species. Leaf mass per area was lower in the growth room than in the field by 43.4–54% and 5.7–21.2% for grasses of high spreading capacity and for non-invasive C4 species, respectively. Little or no response was observed in the proportion of epidermis and mesophyll, but the proportional area of veins plus sclerenchyma was greater in the field than in the growth room for the invasive C4 Sorghum halepense, and the spreading C3 Bromus inermis and Calamagrostis epigeios, while it did not differ for the two non-invasive C4 grasses and the invasive C4 Cynodon dactylon. Leaf intervenial distance was invariant for C4 grasses (except for the non-invasive Chrysopogon gryllus) and the C3 C. epigeios, but changed by 25.1% for the C3 B. inermis. These results suggest that among C4 grasses invasive species exceed non-invasive ones in the plasticity of leaf coarse structure, but not that of leaf morphology and anatomy. However, leaf structure was not less plastic in invasive C4 than in expanding C3 grasses except for intervenial distance.

Tidal marshes as energy sources for commercially important nektonic organisms: stable isotope analysis

Tidal marshes provide nursery habitats for many commercial nektonic species; thus, determining trophic linkages between tidal marshes and aquatic consumers is important for sustain- ing fishery production in estuarine ecosystems. We examined stable isotopes (δ 13 C, δ 15 N) in 4 com- mercial nekton species (Chelon haematocheilus, Synechogobius ommaturus, Lateolabrax maculatus and Exopalaemon carinicauda) in the tidal marshes of the Yangtze River estuary, China. We esti- mated the frequency and range of potential contribution (0 to 100%) from different food sources (benthic microalgae, suspended particulate organic matter, the invasive C4 plant Spartina alterniflora and native C3 plants Phragmites australis and Scirpus mariqueter) to the nektonic consumers, and then pooled the contributions for primary producers with similar isotope values (giving 3 groups: microalgae, invasive C4 plant and native C3 plants). Marsh vascular plants and microalgae were at the base of the food web supporting these nektonic species. For C. haematocheilus and S. ommatu- rus, vascular plants constituted a larger fraction of their carbon source than microalgae. S. alterniflora contributed more than 50% of their total organic carbon and was more important than the native C3 plants. For L. maculatus and E. carinicauda, intermediate δ 13 C values precluded definitive assign- ment of a major carbon source. We have shown that tidal marshes provide important food sources for some dominant estuarine nektonic species, and that the exotic plant S. alterniflora has been incorpo- rated into aquatic food webs of the Yangtze River estuary.

Climatic variability alters the outcome of long‐term community assembly

Summary We used seed additions to test experimentally whether long‐term community assembly unfolds consistently (deterministic model) or whether different outcomes are possible depending on variations in climate and the presence of exotic dominants (stochastic model). The experiment was conducted in homogeneous semi‐arid grassland on the northern Great Plains of North America. Native grass seed additions in each of 3 years (1994–96) were combined factorially with repeated selective herbicide applications to reduce the cover of the extant dominant, a Eurasian C3 grass (Agropyron cristatum). We assessed whether composition converged or diverged among treatments between 1996 and 2004. The outcome of long‐term community assembly varied by establishment year, in association with significant variations in monthly growing‐season precipitation from 68‐year averages during the years when seed was added. In 2004, the added native C4 grass Bouteloua gracilis dominated plots sprayed and seeded in 1994 and 1995, resisting re‐invasion and having significantly higher plot diversity. In contrast, plots sown in 1996 – one of the driest years on record – reverted to the exotic C3 grass. Seeded but unsprayed plots maintained their pre‐treatment dominance by exotic grass. Rainfall variability was also associated with the extent of exotic grass cover and bare soil in unsprayed plots, and natural recruitment by native species from nearby prairie. Establishment success varied significantly among the added species. All seeded grasses, other than B. gracilis, were mostly absent by 2004. The failure of these native C3 species to establish appears consistent with deterministic assembly models where local abiotic conditions filter out unsuitable species. Synthesis. Our results indicate that stochastic and deterministic processes operate simultaneously to influence community assembly, depending on interactions among climate, seed availability, species identity and disturbance during the initial stages of establishment. Multiple assembly trajectories developed, but the assembled communities did not include all the added species. Conversion of degraded grassland back to native‐dominated grassland was possible, but only with the removal of the exotic dominant and seed additions during years that were suitably wet for establishment. Once formed, however, the assembled native community resisted re‐invasion. This suggests that native grassland restoration will depend on establishment measures as intense as those used during initial cultivation almost a century earlier.

Profiling the Enzymatic Properties and Inhibition of Human Complement Factor B

Human complement factor B is the crucial catalytic component of the C3 convertase enzyme that activates the alternative pathway of complement-mediated immunity. Although a serine protease in its own right, factor B circulates in human serum as an inactive zymogen and there is a crystal structure only for the inactive state of factor B and various fragments. To provide greater insight to the catalytic function and properties of factor B, we have used short para-nitroanilide derivatives of 4- to 15-residue peptides as substrates to profile the catalytic properties of factor B. Among factors found to influence catalytic activity of factor B was an unusual dependence on pH. Non-physiological alkaline conditions strongly promoted substrate cleavage by factor B, consistent with a pH-accessible conformation of the enzyme that may be critical for catalytic function. Small N-terminal extensions to conventional hexapeptide para-nitroanilide substrates significantly increased catalytic activity of factor B, which was more selective for its cleavage site than trypsin. The new chromogenic assay enabled optimization of catalysis conditions, the profiling of different substrate sequences, and the development of the first reversible and competitive substrate-based inhibitor of factor B. The inhibitor was also shown to prevent in vitro formation of C3a from C3 by factor B, by synthetic and by natural C3 convertase of the alternative complement activation pathway, and to block formation of membrane attack complex. The availability of a reversible substrate-based inhibitor that could stabilize the active conformation of factor B, in conjunction with a pH-promoted higher processing activity, may offer a new avenue to obtain crystal structures of factor B and C3 convertase in an active conformation.

Live Cell Imaging of Outward and Inward Vesiculation Induced by the Complement C5b-9 Complex

Cells resist death induced by the complement membrane attack complex (MAC, C5b-9) by removal of the MAC from their surface by an outward and/or inward vesiculation. To gain an insight into the route of MAC removal, human C9 was tagged with Alexa Fluor 488 and traced within live cells. Tagged C9-AF488 was active in lysis of erythrocytes and K562 cells. Upon treatment of K562 cells with antibody and human serum containing C9-AF488, C9-AF488 containing MAC bound to the cells. Within 5-10 min, the cells started shedding C5b-9-loaded vesicles (0.05-1 mum) by outward vesiculation. Concomitantly, C9-AF488 entered the cells and accumulated in a perinuclear, late recycling compartment, co-localized with endocytosed transferrin-Texas Red. Similar results were obtained with fixed cells in which the MAC was labeled with antibodies directed to a C5b-9 neoepitope. Inhibition of protein kinase C reduced endocytosis of C5b-9. Kinetic analysis demonstrated that peripheral, trypsin-sensitive C5b-9 was cleared from cells at a slower rate relative to fully inserted, trypsin-resistant C5b-9. MAC formation is controlled by CD59, a ubiquitously expressed membrane complement regulator. Analysis at a cell population level showed that the amount of C5b-9-AF488 bound to K562 cells after complement activation was highly heterogeneous and inversely correlated with the CD59 level of expression. Efficient C9-AF488 vesiculation was observed in cells expressing low CD59 levels, suggesting that the protective impact of MAC elimination by vesiculation increases as the level of expression of CD59 decreases.

Activation of C5 via the lectin pathway of complement (B181)

Abstract Activation of C5 via the lectin pathway of complement was analyzed by examining C5 cleaving properties of lectin pathway C3/C5 convertases. C4b-dependent C3/C5 convertase, ZymM1,C4b,C2a and EManM1C4b,C2a (M1 represents MBL-MASPs) assembled on zymosan or sheep erythrocytes coated with mannan (EMan) exhibited a weak affinity for C5 (Km = 2.7–5.3 μM). Very high affinity C5 convertases (ZymM1,C3bC4b,C2a and EManM1,C3bC4b,C2a) were generated when low affinity C4b-dependent C3/C5 convertases were allowed to deposit C3b by cleaving native C3. ZymM1,C3bC4b,C2a and EManM1,C3bC4b,C2a exhibited Km (4.9–8.3 nM) well below the normal concentration of C5 in blood (0.37 μM). Although, the Km, Vmax, and kcat of C3/C5 convertases of lectin pathway were similar to those reported for classical pathway, the ability of C4b to bind C2 indicated that every C4b deposited on zymosan/EMan was capable of forming a convertase. The findings differ from those reported for classical pathway C3/C5 convertase where one out of four C4b molecules deposited formed an enzyme. The four times greater potential of the lectin pathway than the classical pathway in generating C3/C5 convertases and producing proinflammatory products, such as C3a, C4a, and C5a implies that activation of complement via the lectin pathway might be a more prominent contributor to the pathology of inflammatory reactions. Research supported by NIH grant HL073804

A Bacterial Anticomplement

IMMUNOLOGY The complement system generates a finely regulated, yet potent antimicrobial response, making it an attractive target for bacterial virulence factors. Most commonly, endogenous regulatory proteins of the complement system are usurped to switch off complement activation, but the widespread human pathogen Staphylococcus aureus can inactivate the complement cascade by a more direct means. Previous work has shown that the extracellular fibrinogen-binding protein (Efb-C) generated by S. aureus blocks the complement pathway by binding to the thioester-containing domain of the complement C3b protein; indeed, S. aureus strains that lack Efb-C display reduced virulence. Hammel et al. resolve the crystal structures for the C3-binding domain of Efb-C in its unbound state and in complex with the C3d domain of C3 (shown at right). Structure-based functional studies suggest that native C3 is bound by Efb-C in a way that alters its conformation. As a consequence, conversion to C3b is prevented, and participation in the subsequent activation of the complement cascade is also blocked. As well as binding native C3, Efb-C also had high affinity for C3b, again appearing to induce conformational changes, this time in the already activated form of the complement component. Effective targeting of the interface between Efb-C and the C3d domain by a small molecule could be useful in the treatment of S. aureus infection. — SJS Nat. Immunol . 10.1038/ni1450 (2007).

Support for a trimeric collar of myosin binding protein C in cardiac and fast, but not slow, skeletal muscle

Myosin binding protein C (MyBPC) has been shown to have both a structural and regulatory function in the striated muscle sarcomere. We have previously proposed a quaternary structure for this molecule within the sarcomere in which MyBPC takes on a trimeric collar arrangement around the thick filament backbone, based on interactions between domains C5 and C8. We set out to determine whether the C5:C8 interaction is directed by the cardiac-specific insertion in domain C5, and, if not, whether C5:C8 interactions also occur in skeletal MyBPC isoforms, using qualitative and quantitative yeast two-hybrid protein–protein interaction assays. Findings were confirmed by surface plasmon resonance. We demonstrate enhanced interaction between cardiac domain C8 and a cardiac domain C5 from which the insertion loop had been deleted (Ka=1.3×105), as compared to interaction of domain C8 with native cardiac C5 (Ka=8.2×104). Furthermore, we find evidence for a C5:C8 interaction in the fast skeletal isoform (Ka=5.7×105), but not in the slow skeletal isoform. These results indicate that the cardiac-specific insertion does not drive the C5:C8 interaction first demonstrated for the cardiac isoform. Moreover, the finding that the interaction holds true for fast, but not slow, skeletal muscle, may indicate that the trimeric collar arrangement of MyBPC is significant for the regulation of high rates of contraction and relaxation in fast skeletal and cardiac muscle. Further studies are required to explore the quaternary structure of MyBPC in slow skeletal muscle.

A structural basis for complement inhibition by Staphylococcus aureus

To provide insight into bacterial suppression of complement-mediated immunity, we present here structures of a bacterial complement inhibitory protein, both free and bound to its complement target. The 1.25-A structure of the complement component C3-inhibitory domain of Staphylococcus aureus extracellular fibrinogen-binding protein (Efb-C) demonstrated a helical motif involved in complement regulation, whereas the 2.2-A structure of Efb-C bound to the C3d domain of human C3 allowed insight into the recognition of complement proteins by invading pathogens. Our structure-function studies provided evidence for a previously unrecognized mode of complement inhibition whereby Efb-C binds to native C3 and alters the solution conformation of C3 in a way that renders it unable to participate in successful 'downstream' activation of the complement response.

Stable carbon isotope signature of ancient maize agriculture in the soils of Motul de San José, Guatemala

AbstractSoil profiles collected from a 2.5‐km transect radiating from the Maya center of Motul de San José were analyzed for the stable carbon‐isotope composition of their soil organic matter. The residues of maize (Zea mays), the only C4 plant known to have been cultivated in this area by the ancient Maya, impart a carbon‐isotope signature to the underlying soil organic matter reservoir that is distinct from that produced by the native C3 forest vegetation. The varying turnover rates of the humic acid and humin fractions of the soil organic matter allowed us to distinguish between the presence of modern and ancient maize residues in these soils, and to delineate the lateral extent of maize cultivation at this ancient Maya site. The strongest isotopic evidence of maize residues is preserved in the soils surrounding the peripheral settlement of Chäkokot and at one locality within the urban center of Motul de San José. © 2007 Wiley Periodicals, Inc.

Structure of C3b reveals conformational changes that underlie complement activity

Resistance to infection and clearance of cell debris in mammals depend on the activation of the complement system, which is an important component of innate and adaptive immunity. Central to the complement system is the activated form of C3, called C3b, which attaches covalently to target surfaces to amplify complement response, label cells for phagocytosis and stimulate the adaptive immune response. C3b consists of 1,560 amino-acid residues and has 12 domains. It binds various proteins and receptors to effect its functions. However, it is not known how C3 changes its conformation into C3b and thereby exposes its many binding sites. Here we present the crystal structure at 4-A resolution of the activated complement protein C3b and describe the conformational rearrangements of the 12 domains that take place upon proteolytic activation. In the activated form the thioester is fully exposed for covalent attachment to target surfaces and is more than 85 A away from the buried site in native C3 (ref. 5). Marked domain rearrangements in the alpha-chain present an altered molecular surface, exposing hidden and cryptic sites that are consistent with known putative binding sites of factor B and several complement regulators. The structural data indicate that the large conformational changes in the proteolytic activation and regulation of C3 take place mainly in the first conversion step, from C3 to C3b. These insights are important for the development of strategies to treat immune disorders that involve complement-mediated inflammation.

Lectin Pathway of Bony Fish Complement: Identification of Two Homologs of the Mannose-Binding Lectin Associated with MASP2 in the Common Carp (Cyprinus carpio)

The lectin pathway of complement is considered to be the most ancient complement pathway as inferred from identification of ancient homologs of mannose-binding lectin (MBL) and MBL-associated serine proteases (MASPs) in some invertebrates. MBL homologs with galactose selectivity and an MASP3-like sequence also occur in bony fish, linking the evolution of the lectin complement pathway from invertebrates to higher vertebrates. However, these cannot be considered authentic complement components until confirmatory functional evidence is obtained. Here, we report the isolation and characterization of two MBL homologs from a cyprinid teleost, the common carp, Cyprinus carpio. One, designated GalBL, corresponds to the MBL-like molecule with the galactose specificity. The other is an authentic MBL with mannose specificity. Both were found to associate with a serine protease that cleaves native human C4 into C4b but not C4i with a hydrolyzed thioester. Molecular cloning and phylogenetic analysis revealed this C4-activating protease to be carp MASP2, indicating that MASP2 arose before the emergence of bony fish. Database mining of MBL-like genes reveals that MBL and GalBL genes are arranged in tandem in the zebrafish genome and that both lectins are conserved in the distantly related puffer fish. These results imply that bony fish have developed a diverged set of MBL homologs that function in the lectin complement pathway.

Ecophysiology of Two Native Invasive Woody Species and Two Dominant Warm‐Season Grasses in the Semiarid Grasslands of the Nebraska Sandhills

Populations of Pinus ponderosa and Juniperus virginiana are expanding into semiarid Sandhills grasslands in Nebraska. To evaluate the physiological basis of their success, we measured the seasonal course of leaf gas exchange, plant water status, and carbon isotope discrimination in these two native trees and two native C4 grasses (Schizachyrium scoparium and Panicum virgatum). Compared to the trees, grasses had higher net photosynthetic rates (Anet) and water use efficiency (WUE) and more negative predawn and midday water potentials (Ψ) in June and July. While leaf Ψ and rates of leaf gas exchange declined for all four species during August, the Ψmid of the grasses were significantly more negative than those of the two trees. The deeply rooted trees maintained water status during summer, in contrast to the grasses, which senesced. Juniperus virginiana in particular was well adapted to xeric conditions, with low stomatal conductance, high WUE, and positive Anet at low Ψ. The highest values of Anet were observed in May for J. virginiana and in May and September for P. ponderosa. Both species maintained low but positive Anet throughout the winter at temperatures above 0°C. Leaf carbon isotopic signature differed between tree and grass species but did not exhibit significant within‐species seasonal variability. The semiarid grassland climate of Nebraska does not appear to limit P. ponderosa and J. virginiana, which use growth during the nongrowing season and access to deep soil moisture to compensate for growing‐season drought.

Studies on the solvent accessibility of native C3 and its fragments, as analyzed by HDX-MS

Studies on the solvent accessibility of native C3 and its fragments, as analyzed by HDX-MS

Structural insights into the central complement component C3

C3 is a central protein of the complement system, which is important to immune defense and provides a link between innate and adaptive immunity. Three pathways of complement activation converge at the activation of C3 yielding a diverse set of biological responses. This versatile and flexible molecule interacts with various proteins to fulfill its functions. Here we review recent insights gained from the crystal structure determinations of human, native C3 and its physiological down-regulation product C3c. The data provided, for the first time, a complete and detailed view of the composition and arrangement of the domains in C3. Comparison of C3 with C3c indicates marked flexibility of the molecule, particularly in the α-chain. We discuss the observed domain rearrangements, conformational changes and the location of various protein binding sites. These detailed, and structural, insights are important for developing models of the molecular mechanisms underlying the diverse biological activities of this large and complex molecule.