A Host−Guest Set of Triple-Helical Peptides:  Stability of Gly-X-Y Triplets Containing Common Nonpolar Residues

Abstract

Host-guest peptide sets have been useful in evaluating the propensity of different amino acids to adopt an alpha-helical or beta-sheet form, and this concept is applied here to the triple-helical conformation. A set of host-guest peptides of the form acetyl-(Gly-Pro-Hyp)3-Gly-X-Y-(Gly-Pro-Hyp)4-Gly-GlyCONH2 was designed to evaluate the contribution of an isolated Gly-X-Y triplet to triple-helix stability in a defined environment. Peptides were synthesized to include guest triplets with the X and Y positions occupied by the most common nonpolar residues found in collagen: Pro (X position) and Hyp (Y position); Ala; Leu, the most frequent hydrophobic residue; and Phe, the only commonly occurring aromatic residue. The guest triplets of the 12 peptides synthesized represent 35% of the sequence found in the alpha 1 chain of type I collagen. All peptides formed stable triple-helical structures, and the peptides showed a range of thermal stabilities (Tm = 21-44 degrees C), depending on the identity of the guest triplet. Thermodynamic calculations indicate these peptides have a range of free energy values (delta delta G = 9 kcal/mol) and suggest that favorable entropy is the dominant factor in increased stability. Replacement of Ala by Leu in the X position did not affect the thermal stability, while an Ala to Leu change in the Y position was destabilizing. These data provide experimental evidence that hydrophobic residues do not stabilize the triple helical conformation. Although Leu and Phe are found almost exclusively in the X position in collagens, peptides with Leu and Phe in the Y position formed stable triple-helices. This supports the hypothesis that the X positional preference of these residues relates to their increased potential for intermolecular hydrophobic interactions rather than their destabilization of the triple-helical molecule. These studies establish the utility of host-guest peptides in defining a scale of triple-helix propensities and in clarifying the interactions stabilizing the triple-helical conformation.

Integrin Activation State Determines Selectivity for Novel Recognition Sites in Fibrillar Collagens

“…All the double triplets were synthesized in two different flanking hosts (22,23), (GPP) 5 and (GPO) 3 , which provide the minimal structure to maintain triple helix stability at 20°C (24). We identified new sequences analogous to GFOGER at conserved loci within the collagen helix that can bind integrin, and confirmed the reactivity of GLOGER and GASGER.…”

Destabilization of osteogenesis imperfecta collagen-like model peptides correlates with the identity of the residue replacing glycine

“…The influence of those residues replacing Gly in OI mutations is reported herein for a host-guest peptide model. The host peptide consists of eight Gly-Pro-Hyp triplets, a model of the most stable collagenous environment (19). Near the center, one Gly residue is replaced with Ala, Arg, Asp, Cys, Glu, Ser, or Val, and the effect on stability is analyzed.…”

Characterization of a human surfactant protein A1 (SP-A1) gene-specific antibody; SP-A1 content variation among individuals of varying age and pulmonary health

“…SP-A1 has a Cys in the Y position (GEC) referred to as Cys 85 , whereas the SP-A2 has an Arg (GER) referred to as Arg 85 . Studies of host/guest triple helical peptides, where a single guest Gly-X-Y with substitutions in X and Y positions was studied while embedded in a collagen host peptide, showed that an Arg in the Y position associates with high stability of the host/guest peptide (9,61,73). Based on these studies, we have previously proposed (21,22) that Arg 85 (SP-A2) has a higher local microstability compared with Cys 85 (SP-A1).…”

“…1. The 21-aa peptide contained three SP-A1 gene-specific aa: Asp 73 , Ile 81 , and Cys 85 at aa positions 73,81, and 85 of SP-A1. The proline residues at positions 76, 79, and 82 are either hydroxylated or unhydroxylated.…”

“…However, at the mRNA level, the SP-A1-to-SP-A2 ratio has been shown to vary considerably (39). Although the SP-A1 and SP-A2 variants differ collectively at 10 aa, the gene specificity between SP-A1 and SP-A2 variants is determined by differences in 4 aa (66,73,81,85) within the collagen region. Of the 10 aa differences between SP-A1 and SP-A2 variants (10,16,40), 2 aa are located in the NH 2 -terminal signal peptide, 6 in the collagenlike region, and 2 in the carbohydrate recognition domain.…”