Min-21 and min-23, the smallest peptides that fold like a cystine-stabilized beta-sheet motif: design, solution structure, and thermal stability.
Small disulfide-rich proteins provide examples of simple and stable scaffolds for design purposes. The cystine-stabilized beta-sheet (CSB) motif is one such elementary structural motif and is found in many protein families with no evolutionary relationships. In this paper, we present NMR structural studies and stability measurements of two short peptides of 21 and 23 residues that correspond to the isolated CSB motif taken from a 28-residue squash trypsin inhibitor. The two peptides contain two disulfide bridges instead of three for the parent protein, but were shown to fold in a native-like fashion, indicating that the CSB motif can be considered an autonomous folding unit. The 23-residue peptide was truncated at the N-terminus. It has a well-defined conformation close to that of the parent squash inhibitor, and although less stable than the native protein, it still exhibits a high T(m) of about 100 degrees C. We suggest that this peptide is a very good starting building block for engineering new bioactive molecules by grafting different active or recognition sites onto it. The 21-residue peptide was further shortened by removing two residues in the loop connecting the second and third cysteines. This peptide exhibited a less well-defined conformation and is less stable by about 1 kcal mol(-)(1), but it might be useful if a higher flexibility is desired. The lower stability of the 21-residue peptide is supposed to result from inadequate lengths of segments connecting the first three cysteines, thus providing new insights into the structural determinants of the CSB motif.
Heitz
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A
Centre de Biochimie Structurale, Faculté de pharmacie, UMR9955 CNRS-INSERM-Université Montpellier I, France.
Le-Nguyen
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D
Chiche
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L
D000595:Amino Acid Sequence; D019662:Cucurbitaceae; D003553:Cystine / Q000737:chemistry*; D004220:Disulfides / Q000737:chemistry; D006358:Hot Temperature; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D010455:Peptides / Q000138:chemical synthesis* / Q000737:chemistry* / Q000378:metabolism; D010940:Plant Proteins; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017386:Sequence Homology, Amino Acid; D012996:Solutions; D013816:Thermodynamics; D014361:Trypsin Inhibitors / Q000737:chemistry*
D004220:Disulfides; D010455:Peptides; D010940:Plant Proteins; D012996:Solutions; D014361:Trypsin Inhibitors; C063950:trypsin inhibitor EETI II protein, plant; D003553:Cystine