Synthesis, purification and initial structural characterization of octarellin, a de novo polypeptide modelled on the alpha/beta-barrel proteins.
We have attempted to construct an artificial polypeptide that folds like the eight-stranded parallel beta-barrel structures. Our approach consists of repeating eight times a unit peptide designed to adopt a 'beta-strand/alpha-helix' pattern. A first 'test' sequence for this structural unit was deduced from a series of parameters defined after an analysis of three natural alpha/beta-barrel proteins and including principally the lengths of the secondary structure elements, the alpha/beta packing and the fitting on average Garnier profiles. The gene encoding this structural unit was synthesized, cloned and expressed in Escherichia coli either as a monomer or as direct repeats of 2-12 units. Preliminary structural characterization of the 7-, 8- and 9-fold unit polypeptides by circular dichroism measurements indicates the presence of the predicted amount of alpha-helix in the three proteins. Further analysis by urea-gradient gel electrophoresis demonstrates that, in the conditions tested, only the 8-fold unit polypeptide forms a compact structure through a cooperative and rapid two-state folding transition involving long-range molecular interactions.
Université de Liège, Institut de Chimie, Sart Tilman, Belgium.
K
K
Goraj
A
A
Renard
J A
JA
Martial
D000595:Amino Acid Sequence; D001483:Base Sequence; D002942:Circular Dichroism; D003001:Cloning, Molecular; D001330:Electronic Data Processing; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli; D008969:Molecular Sequence Data; D010455:Peptides / Q000235:genetics; D010957:Plasmids; D011487:Protein Conformation; D015202:Protein Engineering
D010455:Peptides