Abstract
An approach is described for the de novo design of protein-like structures in which synthetic combinatorial libraries (SCLs) were incorporated into an amphipathic alpha-helical scaffold (an 18-mer sequence made up of leucine and lysine residues) to generate conformationally defined SCLs. In particular, the SCLs in which the "combinatorialized" positions were on the hydrophilic face showed an alpha-helical conformation in mild buffer. These SCLs were used to generate context-independent but position-dependent scales of alpha-helical propensity for the L-amino acids. These scales were then used to design highly alpha-helical peptides that self-associated in mild buffer. The same approach was also found to permit the identification of conformation-dependent decarboxylation catalysts.
Highlights
In an initial effort directed toward the design of new proteinlike structures using synthetic combinatorial libraries (SCLs) approaches, conformationally defined combinatorial libraries have been designed by randomizing five positions in two different 26-amino acid-long naturally occurring peptides
The structural nature of these libraries was used to study the helical propensity of amino acids, which permitted the identification of peptides that self-aggregated into ␣-helical conformations in mild buffer
Design of Conformationally Defined SCLs—The 18-mer peptide scaffold used in these studies (Table I) was found in earlier studies to be a random coil in mild buffer but to adopt a monomeric amphipathic ␣-helical conformation [6] both in trifluoroethanol, a solvent known to promote ␣-helical conformation [19], and in lipid environment
Summary
SCLs and Individual Peptide Synthesis—The SCLs and individual peptides were prepared by simultaneous multiple peptide synthesis using t-Boc chemistry as described elsewhere [13]. Final cleavage and deprotection steps were carried out using a “lowhigh” hydrogen fluoride procedure [15, 16]. Individual peptides were purified by preparative reversed phase-high performance liquid chromatography (HPLC) using a DeltaPrep 3000 reversed phase-HPLC combined with a Foxy fraction collector (Millipore, Waters Division, San Francisco, CA). Analytical reversed phase-HPLC and laser desorption time-of-flight mass spectroscopy (Kompact Maldi-Tof mass spectrometer, Kratos, Ramsey, NJ) were used to determine the purity and identity of the individual peptides. Circular Dichroism Measurements—All measurements were carried out on a Jasco J-720 CD spectropolarimeter (Eaton, MD) in conjunction with a Neslab RTE 110 water bath and temperature controller (Dublin, CA). CD spectra were the average of a series of 3–7 scans made at.
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