Crystallization and preliminary X-ray diffraction analysis of eukaryotic α2 -macroglobulin family members modified by methylamine, proteases and glycosidases.

α2 -Macroglobulin (α2 M) has many functions in vertebrate physiology. To understand the basis of such functions, high-resolution structural models of its conformations and complexes with interacting partners are required. In an attempt to grow crystals that diffract to high or medium resolution, we isolated native human α2 M (hα2 M) and its counterpart from chicken egg white (ovostatin) from natural sources.

Purification of human complement protein C5.

Complement C5 is cleaved by proteolysis in the terminal phase of complement activation generating the pro-inflammatory C5a and membrane attack complex nucleator C5b. Whereas purification of its paralogues C3 and C4 from plasma is relatively straightforward, C5 purification is more complicated due to the lower amounts present and overlaps with the much more abundant C3 during several chromatographic steps. Here we describe our procedure for purifying homogenous, monodisperse, and crystallizable C5.

Structural basis for activation of the complement system by component C4 cleavage.

An essential aspect of innate immunity is recognition of molecular patterns on the surface of pathogens or altered self through the lectin and classical pathways, two of the three well-established activation pathways of the complement system. This recognition causes activation of the MASP-2 or the C1s serine proteases followed by cleavage of the protein C4. Here we present the crystal structures of the 203-kDa human C4 and the 245-kDa C4·MASP-2 substrate·enzyme complex.

The crystal structure of human α2-macroglobulin reveals a unique molecular cage.

I'm your Venus: the crystal structure of the human methylamine-induced form of α(2)-macroglobulin (α(2)M) shows its large central cavity can accommodate two medium-sized proteinases. Twelve major entrances provide access for small substrates to the cavity and the still-active trapped "prey". The structure unveils the molecular basis of the unique "venus flytrap" mechanism of α(2)M.

Substrate recognition by complement convertases revealed in the C5-cobra venom factor complex.

Complement acts as a danger-sensing system in the innate immune system, and its activation initiates a strong inflammatory response and cleavage of the proteins C3 and C5 by proteolytic enzymes, the convertases. These contain a non-catalytic substrate contacting subunit (C3b or C4b) in complex with a protease subunit (Bb or C2a). We determined the crystal structures of the C3b homologue cobra venom factor (CVF) in complex with C5, and in complex with C5 and the inhibitor SSL7 at 4.