ABSTRACT
A G protein-coupled receptor (GPCR) agonist protein, thaumatin, was site-specifically conjugated at the N- or C-terminus with a fluorophore for visualization of GPCR:agonist interactions. The N-terminus was specifically conjugated using a synthetic 2-pyridinecarboxyaldehyde reagent. The interaction profiles observed for N- and C-terminal conjugates were varied; N-terminal conjugates interacted very weakly with the GPCR of interest, whereas C-terminal conjugates bound to the receptor. These chemical biology tools allow interactions of therapeutic proteins:GPCR to be monitored and visualized. The methodology used for site-specific bioconjugation represents an advance in application of 2-pyridinecarboxyaldehydes for N-terminal specific bioconjugations.
Subject(s)
Plant Proteins/chemistry , Plant Proteins/pharmacology , Receptors, G-Protein-Coupled/agonists , Sweetening Agents/chemistry , Sweetening Agents/pharmacology , Animals , Cell Line , Drug Design , Fluorescence , Fluorescent Dyes/chemistry , Humans , Microscopy, Fluorescence/methods , Optical Imaging , Protein Binding , Receptors, G-Protein-Coupled/metabolism , Recombinant Proteins/chemistry , Recombinant Proteins/pharmacologyABSTRACT
IMPORTANCE: In youth, facial aesthetic units flow together without perceptible division. The face appears as a single dynamic structure with a smooth contour and very little if any shadowing between different anatomical regions. As one ages, facial aesthetic units slowly become distinct. This process may be a consequence of differences in skin thickness, composition of subcutaneous tissue, contour of the facial skeleton, and location of facial ligaments. Although the impact of aesthetic unit separation is clinically apparent, its fundamental role in perceived facial aging has not yet been defined empirically. OBJECTIVES: To evaluate and define the effect of aesthetic unit separation on facial aging and to empirically validate the rationale for the blending of aesthetic units as a principle for facial rejuvenation. DESIGN, SETTING, AND PARTICIPANTS: We prepared the photographs of 7 women for experimental evaluation of the presence or absence of facial aesthetic unit separation. Photographic stimuli were then presented to 24 naive observers in a blinded paired comparison. For each stimulus pair, observers were asked to select the facial photograph that they considered to be more youthful in appearance. Each stimulus was compared with all others. MAIN OUTCOMES AND MEASURES: We calculated a preference score for the total number of times any photograph was chosen to be more youthful compared with all others. Paired t tests were used to compare the preference scores between the facial stimuli with and without aesthetic unit separation. RESULTS: We generated 4032 responses for analysis. Photographs without facial aesthetic unit separation were consistently judged to be more youthful than their aged original or modified counterparts, with mean preference scores of 0.66 and 0.33, respectively (P ≤ .047). When we selected the paired stimulus that directly compared one photograph with aesthetic unit separation with another with blended aesthetic units (2015 pairs), observers indicated that the photograph with the blended aesthetic unit was younger 95% of the time. Within-rater reliability was found to be very good (r = 0.88). CONCLUSIONS AND RELEVANCE: Our data support the hypothesis that facial aesthetic unit separation influences perceived facial youthfulness among photographs of women. The presence of facial aesthetic unit separation results in a less youthful appearance. Based on these empirical data, the concept of facial aesthetic unit separation appears to play a significant role in perceived facial aging. LEVEL OF EVIDENCE: NA.
