Insights into the role of the histidines in the structure and stability of cytochrome c |
| |
Authors: | Federica Sinibaldi Barry D Howes M Cristina Piro Paola Caroppi Giampiero Mei Franca Ascoli Giulietta Smulevich Roberto Santucci |
| |
Institution: | (1) Dipartimento di Medicina Sperimentale e Scienze Biochimiche, Università di Roma ‘Tor Vergata’, via Montpellier 1, 00133 Rome, Italy;(2) Dipartimento di Chimica, Università degli Studi di Firenze, via della Lastruccia 3, 50019 Sesto Fiorentino (FI), Italy |
| |
Abstract: | In this paper we investigate the role played by each histidine in the amino acid sequence of yeast iso-1-cytochrome c (with the exception of H18, the residue axially coordinated to the heme iron) in determining the protein structure and stability.
To this end, we have generated and characterized the double mutants H26Y/H33Y, H26Y/H39K and H33Y/H39K obtained from the C102T
variant of the protein, which retain only one histidine side chain in the amino acid sequence. In particular, the H39K mutation
inserts a lysine at position 39 as in the sequence of equine cytochrome c. The H26Y/H33Y/H39K triple mutant, which lacks all three histidines, was also produced and its spectroscopic properties are
compared with those of the double mutants. The data highlight the critical role played by H26 in determining protein stability.
Recombinant horse cytochrome c and the corresponding H26Y mutant were also generated and characterized. Since equine cytochrome c exhibits higher stability than the yeast protein, this provides a valuable opportunity to understand the role played by the
invariant H26 residue in determining structure and stability. |
| |
Keywords: | Cytochrome c Site-directed mutagenesis Intermediate state Stability Resonance Raman spectroscopy |
本文献已被 SpringerLink 等数据库收录! |
|