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Defining the minimum size of a hydrophobic cluster in two‐stranded α‐helical coiled‐coils: Effects on protein stability
Author(s) -
Lu Stephen M.,
Hodges Robert S.
Publication year - 2004
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1110/ps.03443204
Subject(s) - coiled coil , crystallography , chemistry , hydrophobic effect , peptide , heptad repeat , protein structure , biophysics , stereochemistry , peptide sequence , biochemistry , biology , gene
The α‐helical coiled‐coil motif is characterized by a heptad repeat pattern ( abcdefg ) n in which residues a and d form the hydrophobic core. Long coiled‐coils (e.g., tropomyosin, 284 residues per polypeptide chain) typically do not have a continuous hydrophobic core of stabilizing residues, but rather one that consists of alternating clusters of stabilizing and destabilizing residues. We have arbitrarily defined a cluster as a minimum of three consecutive stabilizing or destabilizing residues in the hydrophobic core. We report here on a series of two‐stranded, disulfide‐bridged parallel α‐helical coiled‐coils that contain a central cassette of three consecutive hydrophobic core positions ( d, a , and d ) with a destabilizing cluster of three consecutive Ala residues in the hydrophobic core on each side of the cassette. The effect of adding one to three stabilizing hydrophobes in these positions (Leu or Ile; denoted as •) was investigated. Alanine residues (denoted as ○) are used to represent destabilizing residues. The peptide with three Ala residues in the d a d cassette positions (○○○) was among the least stable coiled‐coil (T m = 39.3°C and Urea 1/2 = 1.9 M). Surprisingly, the addition of one stabilizing hydrophobe (Leu) to the cassette or two stabilizing hydrophobes (Leu), still interspersed by an Ala in the cassette (•○•), also did not lead to any gain in stability. However, peptides with two adjacent hydrophobes in the cassette (••○)(○••) did show a gain in stability of 0.9 kcal/mole over the peptide with two interspersed hydrophobes (•○•). Because the latter three peptides have the same inherent hydrophobicity, the juxtaposition of stabilizing hydrophobes leads to a synergistic effect, and thus a clustering effect. The addition of a third stabilizing hydrophobe to the cassette (•••) resulted in a further synergistic gain in stability of 1.7 kcal/mole (T m = 54.1°C and Urea 1/2 = 3.3M). Therefore, the role of hydrophobicity in the hydrophobic core of coiled‐coils is extremely context dependent and clustering is an important aspect of protein folding and stability.