Remodelling the folding of thioredoxin by removal of the C-terminal helix.

JAVIER SANTOS; JOSÉ MARÍA DELFINO

Montpellier, France.

Congreso; 15th. IUPAB and 5th. EBSA International Biophysics Congress.; 2005

IUPAB

E. coli thioredoxin (TRX) is a monomeric a/b protein of 108 amino acids with a fold characterized by a central beta sheet surrounded by alpha helices. Two subdomains are topologically noticeable, but it is unclear whether their folding occurs in a concerted fashion. Subdomain TRX1-73 has been extensively studied as a model of a partially folded, with no tertiary or persistent secondary structure. This work describes the expression and characterization -by circular dichroism (CD), fluorescence emission, size exclusion chromatography,  chemical cross-linking and light scattering- of a novel engineered fragment (TRX1-93) lacking the last stretch of 15 amino acids. After refolding from inclusion bodies, TRX1-93 shows a strong propensity to form soluble oligomers endowed with distinctive optical properties unlike those observed for the full protein. Although TRX1-93 also shows significant changes in secondary structure, Trp residues appear to occupy rigid and apolar environments. These findings support the existence of an alternatively folded form for TRX1-93. In addition, the secondary structure content of chemically synthesized peptide TRX94-108 and its ability to complement fragment TRX 1-93 upon refolding were also evaluated by CD. Taken together, the data herein presented shed light upon issues such as the distribution of information content relevant for folding along the polypeptide chain in regard to conformational stability. With grants from ANPCyT, UBACyT and CONICET.