Abstract
Three mutants of the anaphylatoxin C5a were prepared with positions 2, 64 and 70, respectively, substituted by tryptophan. The last mutant was additionally labelled at Cys27 for fluorescence energy transfer (FET) measurements. The structural integrity and biological activity of the molecules were not affected. Fluorescence anisotropy decay (FAD) measurements showed that the rotational correlation time for tryptophan decreases in the order: [Trp2]rhC5a > [Trp64]rhC5a > [Trp70]rhC5a, indicating an increasing mobility of the side chain. Measurements of the fluorescence energy transfer from Trp70 to the 1,5-AEDANS group at Cys27 yielded a distance distribution of 2.4 ± 0.8 nm. This value is compatible with the C-terminal chain being arranged as a slightly stretched helix pointing away from the body of the molecule.
| Original language | English |
|---|---|
| Journal | Biophysical Chemistry |
| Volume | 46 |
| Issue number | 3 |
| Pages (from-to) | 237-248 |
| Number of pages | 12 |
| ISSN | 0301-4622 |
| DOIs | |
| Publication status | Published - 01.05.1993 |
Funding
Thanks are due to following members of the lnstitut fiir Biochemie: Manfred Dewor for the amino acid analyses, to Jiirgen Stahl for the CD measurements, to Berthold Wroblowski for determining the secondary structural composition of rhC5a on the bases of these results and to Edgar Jacoby for computer graphic distance determinations. Furthermore, we wish to thank Dr. G. Marius Clore for providing the backbone coordinates of the NMR structure of bovine C5a. This work was supported by a grant from the Bun-desminister fiir Forschung und Technologie der Bundesrepublik Deutschland (Fbrderkenn-zeichen 01 VM 8901/4X
