Complement fragment (C)5a is a 74 residue pro-inflammatory polypeptide produced during activation of the complement cascade of serum proteins in response to foreign surfaces such as microorganisms and tissue damaged by physical or chemical injury. C5a binds to at least two seven-transmembrane domain receptors, C5aR (C5R1, CD88) and C5L2 (gpr77), expressed ubiquitously on a wide variety of cells but particularly on the surface of immune cells like macrophages, neutrophils and T cells. C5aR is a classical G protein-coupled receptor that signals through Gai and Ga16, whereas C5L2 does not appear to couple to G proteins and has no known signalling activity. Although C5a was first described as an anaphylatoxin and later as a leukocyte chemoattractant, the widespread expression of C5aR suggested more general functionality. Our understanding of the physiology of C5a has improved significantly in recent years through exploitation of receptor knockout and knockin mice, C5 and C5a antibodies, soluble recombinant C5a and C5a analogues and newly developed receptor antagonists. C5a is now also implicated in non-immunological functions associated with developmental biology, CNS development and neurodegeneration, tissue regeneration, and haematopoiesis. Combined receptor mutagenesis, molecular modelling, structure-activity relationship studies and species dependence for ligand potency on C5aR have been helpful for identifying ligand binding sites on the receptor and for defining mechanisms of receptor activation and inactivation. This review will highlight major developments in C5a receptor research that support C5aR as an important therapeutic target. The intriguing possibilities raised by the existence of a non-signalling C5a receptor are also discussed.

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Parasite resistance to antimalarial drugs is a serious threat to human health, and novel agents that act on enzymes essential for parasite metabolism, such as proteases, are attractive targets for drug development. Recent studies have shown that clinically utilized human immunodeficiency virus (HIV) protease inhibitors can inhibit the in vitro growth of Plasmodium falciparum at or below concentrations found in human plasma after oral drug administration. The most potent in vitro antimalarial effects have been obtained for parasites treated with saquinavir, ritonavir, or lopinavir, findings confirmed in this study for a genetically distinct P. falciparum line (3D7). To investigate the potential in vivo activity of antiretroviral protease inhibitors (ARPIs) against malaria, we examined the effect of ARPI combinations in a murine model of malaria. In mice infected with Plasmodium chabaudi AS and treated orally with ritonavir-saquinavir or ritonavir-lopinavir, a delay in patency and a significant attenuation of parasitemia were observed. Using modeling and ligand docking studies we examined putative ligand binding sites of ARPIs in aspartyl proteases of P. falciparum (plasmepsins II and IV) and P. chabaudi (plasmepsin) and found that these in silico analyses support the antimalarial activity hypothesized to be mediated through inhibition of these enzymes. In addition, in vitro enzyme assays demonstrated that P. falciparum plasmepsins II and IV are both inhibited by the ARPIs saquinavir, ritonavir, and lopinavir. The combined results suggest that ARPIs have useful antimalarial activity that may be especially relevant in geographical regions where HIV and P. falciparum infections are both endemic.

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28Proteinase-activated receptor 2 (PAR 2 ) is a G protein-coupled receptor involved in 29 metabolism, inflammation, and cancers. It is activated by proteolysis, which exposes a 30 nascent N-terminal sequence that becomes a tethered agonist. Short synthetic peptides 31 corresponding to this sequence also activate PAR 2 , while small organic molecules 32 show promising PAR 2 antagonism. Developing PAR 2 ligands into pharmaceuticals is 33 hindered by a lack of knowledge of how synthetic ligands interact with and 34 differentially modulate PAR 2 . Guided by PAR 2 homology modeling and ligand 35 docking based on bovine rhodopsin, followed by cross-checking with newer PAR 2 36 models based on ORL-1 and PAR 1 , site-directed mutagenesis of PAR 2 was used to 37 investigate the pharmacology of three agonists (two synthetic agonists and trypsin-38 exposed tethered ligand) and one antagonist for modulation of PAR 2 signaling. 39Effects of 28 PAR 2 mutations were examined for PAR 2 -mediated calcium 40 mobilization and key mutants were selected for measuring ligand binding. Nineteen 41 of twenty-eight PAR 2 mutations reduced the potency of at least one ligand by >10-42 fold. Key residues mapped predominantly to a cluster in the transmembrane (TM) 43 domain of PAR 2 , differentially influence intracellular Ca 2+ induced by synthetic 44 agonists versus a native agonist, and highlight subtly different TM residues involved 45 in receptor activation. This is the first evidence highlighting the importance of the 46 PAR 2 TM region for receptor activation by synthetic PAR 2 agonists and antagonists. 47The trypsin-cleaved N-terminus that activates PAR 2 was unaffected by the same 48 residues as synthetic peptides, challenging the widespread practice of substituting 49 peptides for proteases to characterize PAR 2 physiology. 50 51