Association between N-Terminal Pyrenes Stabilizes the Collagen Triple Helix.

Collagen model peptides featuring the fluorophore pyrene at their N-termini have been synthesized, and their thermal denaturation has been examined using circular dichroism (CD) and fluorescence spectroscopies. Flanking the (Pro-Hyp-Gly)7 core of the peptide monomers at positions 1 and/or 23 in the primary sequence, Lys residues were introduced to ensure water solubility. Triple helices derived from such peptides show a broad excimer emission at ∼480 nm, indicative of interaction between the pyrene units. CD experiments show that the fluorophores enhance helix stability primarily through entropic effects. Unfolding temperatures (Tm) increase by up to 7 °C for systems with N-terminal lysine residues and by up to 21 °C for systems in which the first-position Lys is replaced by Ala. Tm values derived from fluorescence measurements (at 50 µM) typically lie within ∼1 °C of those obtained using CD (at 200 µM). Computational modeling in a water continuum using B3LYP-GD3 and M06-2X functionals predicts that face-to-face association of fluorophores can occur while H-bonding within the [(POG)n]3 assembly is retained. Such parallel stacking is consistent with hydrophobically driven stabilization. Labeling collagen peptides with pyrene is a synthetically simple way to promote triple helicity while providing a means to obtain Tm data on relatively dilute samples.

Social Status-Related Differences in Motor Activity Between Wild-Type and Mutant Zebrafish.

Use of zebrafish as a model organism in biomedical research has led to the generation of many genetically modified mutant lines to investigate various aspects of developmental and cellular processes. However, the broader effects of the underlying mutations on social and motor behavior remain poorly examined. Here, we compared the dynamics of social interactions in the Tüpfel long-fin nacre mutant line, which lacks skin pigmentation, to wild-type zebrafish; and we determined whether status-dependent differences in escape and swimming behavior existed within each strain. We show that despite similarities in aggressive activity, Tüpfel long-fin nacre pairs exhibit unstable social relationships characterized by frequent reversals in social dominance compared to wild-type pairs. The lack of strong dominance relationships in Tüpfel long-fin nacre pairs correlates with weak territoriality and overlapping spatial distribution of dominants and subordinates. Conversely, wild-type dominants displayed strong territoriality that severely limited the movement of subordinates. Additionally, the sensitivity of the startle escape response was significantly higher in wild-type subordinates compared to dominants. However, status-related differences in sensitivity of escape response in Tüpfel long-fin nacre pairs were absent. Finally, we present evidence suggesting that these differences could be a consequence of a disruption of proper visual social signals. We show that in wild-type pairs dominants are more conspicuous, and that in wild-type and Tüpfel long-fin nacre pairings wild-type fish are more likely to dominate Tüpfel long-fin nacres. Our results serve as a cautionary note in research design when morphologically engineered zebrafish for color differences are utilized in the study of social behavior and central nervous system function.