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Chemical synthesis of mouse cripto CFC variants
Author(s) -
Marasco Daniela,
Saporito Angela,
Ponticelli Salvatore,
Chambery Angela,
De Falco Sandro,
Pedone Carlo,
Minchiotti Gabriella,
Ruvo Menotti
Publication year - 2006
Publication title -
proteins: structure, function, and bioinformatics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.699
H-Index - 191
eISSN - 1097-0134
pISSN - 0887-3585
DOI - 10.1002/prot.21043
Subject(s) - circular dichroism , chemistry , enzyme , biophysics , kinetics , disulfide bond , peptide , stereochemistry , biochemistry , biology , physics , quantum mechanics
We report for the first time the chemical synthesis of refolded CFC domain of mouse Cripto (mCFC) and of two variants bearing mutations on residues W107 and H104 involved in Alk4 binding. The domains undergo spontaneous and quantitative refolding in about 4 h, yet with very different kinetics. Disulfide linkages have been assessed by enzyme digestion and mass spectrometry analysis of resulting fragments, and the first experimental studies on structural organization have been conducted by circular dichroism spectroscopy under different pH conditions. Upon refolding, the domains considerably change their conformations, although they do not assume canonical structures, and become highly resistant to enzyme degradation. A comparative study of receptor binding shows that the CFC domain can bind Alk4 and confirms the importance of W107 and H104 for receptor recognition. Proteins 2006. © 2006 Wiley‐Liss, Inc.