Primary structure and functional characterization of rat C5a: an anaphylatoxin with unusually high potency.
The anaphylatoxin C5a is a pro-inflammatory factor generated from C5 during complement activation. C5a derived from rat C5 exhibits significantly greater potency compared to C5a from other species. Rat C5a was 25-fold more potent than human C5a for eliciting spasmogenic contraction of guinea pig ileum. Proteolytic removal of the C-terminal arginine of C5a (C5adesArg) reduced spasmogenic potency of rat C5a by only 4-fold compared to a 3,000-fold reduction for human C5adesArg. In addition, rat C5adesArg was 50-fold more potent than human C5adesArg in a guinea pig vascular permeability (in vivo) assay and as a chemotactic factor for human neutrophils. C5a and C5adesArg were purified from zymosan-activated rat serum. Rat C5a, like human C5a, is glycosylated but contains 77 amino acid residues instead of the 74 residues of human C5a. Comparison of the primary structures of rat and human C5a indicated differences at 30 positions including an insert of 3 residues (LLH) in the rat molecule between residue positions 3 and 4 in human C5a. Insertion of residues LLH between Gln-3 and Lys-4 in a recombinant human C5a molecule using site-directed mutagenesis failed to enhance potency. Synthetic C-terminal analogues of rat C5a proved to be measurably more potent than the corresponding human C5a analogues (Ember JA et al., 1993, Protein Sci 2(Suppl 1):159 [Abstr]). We conclude that multiple sequence differences in the C-terminal effector portion and/or elsewhere in rat C5a, but not the LLH insert, account for the significant enhancement in potency of rat C5a over C5a from other species.
Cui
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Institute of Basic Medical Science, Chinese Academy of Medical Sciences, Beijing.
Carney
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DF
Hugli
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D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002633:Chemotaxis; D002850:Chromatography, Gel; D015936:Complement C5a / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology*; D006031:Glycosylation; D006168:Guinea Pigs; D006801:Humans; D007082:Ileum / Q000502:physiology; D008969:Molecular Sequence Data; D009119:Muscle Contraction; D016297:Mutagenesis, Site-Directed; D051381:Rats; D013329:Structure-Activity Relationship
D000596:Amino Acids; D015936:Complement C5a