Malarial vitamin B6 biosynthesis

Abstract

Activation of the mammalian complement cascade induces inflammatory responses and marks pathogens and altered host cells for immune clearance. Central to complement activation is the formation of short-lived protease complexes called convertases. The C3 convertases catalyze the conversion of C3 to its activated form C3b near target surfaces, resulting in opsonisation of target cells and stimulation of the adaptive immune response. By interacting with newly generated C3b molecules, C3 convertases can change their substrate specificity, resulting in the cleavage of C5 to C5b. This process releases the potent anaphylatoxin C5a and activates the terminal pathway of complement, responsible for formation of lytic pores (membrane-attack complexes, MAC) on invading pathogens.The interplay between formation of large multi-domain enzymes, extensive conformational changes and specific proteolytic cleavages highlights the complexity of the complement system.Our investigation of the structural biology of the complement system allowed us to elucidate two new crucial steps of complement activation and regulation: the structures of the pro-convertase of the alternative pathway C3bB and C3bBD*, and the first assembly intermediate of the terminal pathway of complement, C5b6.The structures of the pro-convertase complexes highlight the equilibrium between an initial (closed/loading) state and a subsequent (open/activation) state of the proenzyme factor B bound to C3b. In the transient C3bBD* complex, we show how the open state of FB provides a docking platform for the rate-limiting protease factor D distant from its catalytic site, which has been caught in an activated conformation [1].The C5b6 complex shows marked conformational changes in the conversion from C5 to C5b, highlighting the ability of C6 to capture a labile binding site in C5b to form an assembly competent state provides the first checkpoint in MAC formation. By means of a fruitful collaboration with the laboratories of S.M. Lea in Oxford and O. Llorca in Madrid, we combined our structural data with the electron microscopy reconstruction of the so-called soluble MAC complex (an off-target assembly of the MAC in solution stabilized by specific chaperones). Taken together, these data show how the MAC is assembled and regulated in blood, providing a framework for understanding the role of complement in microbial infection and inflammatory disorders [2].[1]Forneris, F., Ricklin, D., Wu, J., Tzekou, A., Wallace, R.S., Lambris, J.D., Gros, P. (2010 It is perhaps surprising that the main route of vitamin B6 biosynthesis is a recent discovery [1]. The present biochemical research addresses the capability of the enzyme PLP synthase to synthesize vitamin B6 directly from two carbohydrates and the amino acid glutamine [2]. Understanding this enzyme is of relevance in anti-microbial and anti-parasitic drug discovery, particularly in the search for new strategies in anti-malarial therapy [3]. Our initial crystallisation attempts with malarial enzymes did not yield crystals suitabl...