| Abstract |
Since Anfinsens pioneering experiments, it was known that the information necessary to drive protein folding is coded in the linear amino acid sequence of the protein. Despite intense research, the folding process is not fully understood. Various proteins have been used as model systems in order to examine the folding process and numerous theories have tried to interpret the experimental results. A well characterized protein, used in this context, is the Rop protein. Rop is a homodimeric, 4-α-helix bundle, which participates in the control of plasmid copies of the ColE1 family. The topology of each monomer is of the type helix-loop-helix, the loop consisting of three residues. A heptad pattern is present in the Rop sequence and is disturbed only in the loop region. The roles of loop and heptad periodicity in the folding process and stability of Rop have been extensively studied. In this study, we have attempted to farther elucidate the roles of turn and heptad pattern in Rop folding. Towards this aim, four mutants were designed. Two of them comprise altered residues in the turn region, while the other two mutants aim at restoring the heptad periodicity in the same region. A mutant, comprising two proline residues in positions 30 and 31 of wtRop, was constructed. Spectroscopic and thermodynamic studies were carried out and protein crystallization experiments were set up. Results of this work suggest that this mutant is comparable in its spectroscopic and thermodynamic properties to the A31P mutant, which has been compared to a molten-globule like state. This confirms the role of turn residues in the choice of folding pathway. Farther experiments are needed in order to fully characterize this protein. Protein crystals were obtained, but these are not yet suitable for X-ray diffraction experiments. This mutant, although being a loop variant, crystallizes in conditions more close to those of hydrophobic core Rop variants.
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Ο ρόλος της στροφής και του μοτίβου της επτάδας στην αναδίπλωση και σταθερότητα της πρωτεΐνης Rop
