Design and NMR conformational study of a β‐sheet peptide based on Betanova and WW domains

• Published in: Protein science Vol. 15; no. 10; pp. 2278 - 2289
• Main Authors: Fernández‐Escamilla, Ana M., Ventura, Salvador, Serrano, Luis, Jiménez, M. Angeles
• Format: Journal Article
• Language: English
• Published: Bristol Cold Spring Harbor Laboratory Press 01-10-2006
• Subjects: Amino Acid Sequence; antiparallel β‐sheet; Escherichia coli; NMR; Nuclear Magnetic Resonance, Biomolecular; peptide design; peptide structure; Peptides - chemical synthesis; Peptides - chemistry; Protein Conformation; Protein Structure, Secondary; Proteins - chemistry; WW domain
• ISSN: 0961-8368; 1469-896X
• DOI: 10.1110/ps.062186506

A good approach to test our current knowledge on formation of protein β‐sheets is de novo protein design. To obtain a three‐stranded β‐sheet mini‐protein, we have built a series of chimeric peptides by taking as a template a previously designed β‐sheet peptide, Betanova‐LLM, and incorporating N‐ and/or C‐terminal extensions taken from WW domains, the smallest natural β‐sheet domain that is stable in absence of disulfide bridges. Some Betanova‐LLM strand residues were also substituted by those of a prototype WW domain. The designed peptides were cloned and expressed in Escherichia coli. The ability of the purified peptides to adopt β‐sheet structures was examined by circular dichroism (CD). Then, the peptide showing the highest β‐sheet population according to the CD spectra, named 3SBWW‐2, was further investigated by 1H and 13C NMR. Based on NOE and chemical shift data, peptide 3SBWW‐2 adopts a well defined three‐stranded antiparallel β‐sheet structure with a disordered C‐terminal tail. To discern between the contributions to β‐sheet stability of strand residues and the C‐terminal extension, the structural behavior of a control peptide with the same strand residues as 3SBWW‐2 but lacking the C‐terminal extension, named Betanova‐LYYL, was also investigated. β‐Sheet stability in these two peptides, in the parent Betanova‐LLM and in WW‐P, a prototype WW domain, decreased in the order WW‐P > 3SBWW‐2 > Betanova‐LYYL > Betanova‐LLM. Conclusions about the contributions to β‐sheet stability were drawn by comparing structural properties of these four peptides.