Craig A. Gough

The role in cell binding of a beta-bend within the triple helical region in collagen alpha 1 (I) chain: structural and biological evidence for conformational tautomerism on fiber surface.

In its physiological solid state, type I collagen serves as a host for many types of cells. Only the molecules on fiber surface are available for interaction. In this interfacial environment, the conformation of a cell binding domain can be expected to fluctuate between the collagen fold and a distinctive non-collagen molecular marker for recognition and allosteric binding. If the cell binding domain can be localized in contiguous residues within the exposed half of a turn of the triple helix (approximately 15 residues), the need for extensive structural modification and unraveling of the triple helix is avoided. We examined the conformational preferences and biological activity of a synthetic 15-residue peptide (P-15), analogous to the sequence 766GTPGPQGIAGQRGVV780 in the alpha 1 (I) chain. Theoretical studies showed a high potential for a stable beta-bend for the central GIAG sequence. The flanking sequences showed facile transition to extended conformations. Circular dichroism of the synthetic peptide in anisotropic solvents confirmed the presence of beta-strand and beta-bend structures. P-15 inhibited fibroblast binding to collagen in a concentration dependent manner, with near maximal inhibition occurring at a concentration of 7.2 x 10(-6) M. The temporal pattern of cell attachment was altered markedly in the presence of P-15. No inhibition was seen with a peptide P-15(AI), an analogue of P-15 with the central IA residues reversed to AI or with collagen-related peptides (Pro-Pro-Gly)10, (Pro-Hyp-Gly)10, and polyproline, and with several unrelated peptides. Our studies suggest a molecular mechanism for cell binding to collagen fibers based on a conformational transition in collagen molecules on the fiber surface. Since the energy barrier between the collagen fold and beta-strand conformation is low, a local conformational change may be possible in molecules on the fiber surface because of their location in an anisotropic environment. Our observations also suggest that the sequence incorporated in P-15 may be a specific ligand for cells. Unlike other reported cell binding peptides, the residues involved in this interaction are non-polar.

Calculations of the relative free energies of aqueous solvation of several fluorocarbons: A test of the bond potential of mean force correction

The relative free energies of aqueous solvation of several fluorinated derivatives of methane were calculated using the free energy perturbation (FEP) method. The calculations in general duplicated the experimental free energies with relatively good accuracy, but the calculation of the bond potential of mean force (bond‐PMF) contribution [D. A. Pearlman and P. A. Kollman, J. Chem. Phys. 94, 4532 (1991)] was necessary in order to get the most satisfactory agreement with experiment. In particular, it was necessary to use this contribution to obtain even qualitatively correct results for the relative free energies of hydration of methane and tetrafluoromethane. The reasons for this are discussed in terms of the accurate calculation of the effect of the size of the solute. In addition, it is noted that the bond‐PMF contribution is important even for FEP calculations not involving large changes in size, such as the ethanol to ethane perturbation, if the length of a bond to a disappearing atom is changed during...

The Role of Bound Water in the Stability of the Triple-Helical Conformation of (Pro-Pro-Gly)10

There is significant experimental evidence for bound water in collagen and related polymers. (Pro-Pro-Gly)10 [(PPG)10] is a polymer that forms a collagen-like triple-helical structure in aqueous solution. Like collagen, (PPG)10 adopts a structure in which side chains are mostly exposed to solvent, and the backbone polar groups are limited in their ability to form hydrogen bonds with each other. (PPG)10, like collagen, also has many of its backbone polar groups in positions that inhibit complete solvation in aqueous solution; thus the necessity of bound waters for stabilization of the structure. We have constructed a model for bound waters in (PPG)10, based on an examination of the geometry and steric environment of the backbone polar groups. As will become clear, the number of bound waters is determined by the geometry of the backbone carbonyl groups and the steric crowding surrounding them. In this model, each water forms one hydrogen bond with each of two backbone carbonyls from a glycine and a proline in different monomer chains, thus bridging the two carbonyls. The carbonyls in question are quite sterically crowded by neighboring (PPG)10 atoms and would not be likely to experience complete solvation by bulk solvent in aqueous solution. The bound waters are therefore likely to be present even in solution, since otherwise the unsatisfied hydrogen-bonding potential of the carbonyls would destabilize the structure. Other carbonyls also are sterically crowded and possibly prevented from experiencing full solvation, but are not in a favorable geometry for such bridging hydrogen bonds. The intra- and interchain interactions found in a previous computational study of (PPG)10 without bound waters are not disrupted by the addition of waters.

Differential Stability of the Triple Helix of (Pro-Pro-Gly)10 in H2O and D2O: Thermodynamic and Structural Explanations

(Pro-Pro-Gly)10 [(PPG10)], a collagen-like polypeptide, forms a triple-helical, polyproline-II structure in aqueous solution at temperatures somewhat lower than physiological, with a melting temperature of 24.5 degrees C. In this article, we present circular dichroism spectra that demonstrate an increase of the melting temperature with the addition of increasing amounts of D2O to an H2O solution of (PPG)10, with the melting temperature reaching 40 degrees C in pure D2O. A thermodynamic analysis of the data demonstrates that this result is due to an increasing enthalpy of unfolding in D2O vs. H2O. To provide a theoretical explanation for this result, we have used a model for hydration of (PPG)10 that we developed previously, in which inter-chain water bridges are formed between sterically crowded waters and peptide bond carbonyls. Energy minimizations were performed upon this model using hydrogen bond parameters for water, and altered hydrogen bond parameters that reproduced the differences in carbonyl oxygen-water oxygen distances found in small-molecule crystal structures containing oxygen-oxygen hydrogen bonds between organic molecules and H2O or D2O. It was found that using hydrogen bond parameters that reproduced the distance typical of hydrogen bonds to D2O resulted in a significant lowering of the potential energy of hydrated (PPG)10. This lowering of the energy involved energetic terms that were only indirectly related to the altered hydrogen bond parameters, and were therefore not artifactual; the intra-(PPG10) energy, plus the water-(PPG10) van der Waals energy (not including hydrogen bond interactions), were lowered enough to qualitatively account for the lower enthalpy of the triple-helical conformation, relative to the unfolded state, in D2O vs. H2O. This result indicates that the geometry of the carbonyl-D2O hydrogen bonds allows formation of good hydrogen bonds without making as much of an energetic sacrifice from other factors as in the case of hydration by H2O.