Selection of a C5a receptor antagonist from phage libraries attenuating the inflammatory response in immune complex disease and ischemia/reperfusion injury.

A C5a-receptor antagonist was selected from human C5a phage display libraries in which the C terminus of des-Arg74-hC5a was mutated. The selected molecule is a competitive C5a receptor antagonist in vitro and in vivo. Signal transduction is interrupted at the level of G-protein activation.

Analysis of the C5a anaphylatoxin core domain using a C5a phage library selected on differentiated U937 cells.

The human anaphylatoxin C5a is a 74-amino acid comprising polypeptide with a plethora of biological functions. Site directed mutagenesis studies suggest that several residues within the core and the C-terminus mediate the interaction with the C5a receptor. However, the contribution of particular core residues to receptor binding remained to be clarified.

Conservation of the amino-terminal epitope of elongation factor Tu in eubacteria and Archaea.

An epitope of elongation factor Tu (EF-Tu), which is found in organisms in both the bacterial and archaeal domains, was recently defined by mAb 900. To localize the conserved epitope within the EF-Tu molecule and to determine its sequence, SPOTScan analysis of synthetic peptides, Western blot analysis of purified EF-Tu domains and site-directed mutagenesis studies were used. Analysis of mAb 900 binding to overlapping 15-mer peptides encompassing the complete sequence of EF-Tu of Escherichia coli was inconclusive, suggesting three distinct regions may be epitopes.

A detailed analysis of the C5a anaphylatoxin effector domain: selection of C5a phage libraries on differentiated U937 cells.

We have used a phage-display-based system to investigate the effect of simultaneous substitutions within the C5a effector domain. Two different libraries were constructed. In library I, known binding positions 67, 68, 72 and 74 of human complement C5a (hC5a) and in library II, positions 69-73 of hC5a without C-terminal Arg74 (des-Arg74-C5a) were randomly mutated.

Site-directed C3a receptor antibodies from phage display libraries.

Recent cloning of the human C3a receptor (C3aR) revealed that this receptor belongs to the large family of rhodopsin-type receptors. A unique feature of the C3aR is the large second extracellular loop comprising about 175 amino acid residues. We constructed combinatorial phage Ab libraries expressing single chain Fv Abs from BALB/c mice immunized with the affinity-purified second extracellular loop of the C3aR, fused to glutathione-S-transferase.

A selection system to study C5a-C5a-receptor interactions: phage display of a novel C5a anaphylatoxin, Fos-C5aAla27.

Binding and effector domains of the human anaphylatoxin C5a have been determined by either site directed mutagenesis or synthetic peptide studies. However, the lack of specific selection methods, which allow direct investigation of C5a-C5a-receptor interaction made these studies laborious. To overcome these limitations we have constructed a novel Fos-C5a expressed on the tip of a filamentous phage.

Epitope mapping of a C5a neutralizing mAb using a combined approach of phage display, synthetic peptides and site-directed mutagenesis.

Background: The anaphylatoxin C5a is a powerful proinflammatory protein generated on activation of the complement system. Recently, we described an anti-hC5a neoepitope specific mAb, mAb 2925, which was raised against the nonapeptide ISHKDMQLG (C5a-(65-73). This mAb is unique in that it recognizes both hC5a and hC5adesArg, even when it is denatured.

Seroprevalence of immunoglobulin M (IgM) and IgG antibodies to polysaccharides of Streptococcus pneumoniae in different age groups of Ecuadorian and German children.

The age-specific prevalence of serum immunoglobulin M (IgM) antibody to capsular polysaccharides of Streptococcus pneumoniae, as detected by enzyme-linked immunosorbent assay, was studied in 1,301 Ecuadorian children enrolled in a national nutrition and health survey. This prevalence was 6% in infants < 6 months old and increased to 28% in children 6 to 11 months old, 49% in those 12 to 17 months old, and 58% in those 18 to 23 months old. About 80% of the 5-year-old children had this antibody.