Functional Comparison of Human and Zebra Fish FKBP52 Confirms the Importance of the Proline-Rich Loop for Regulation of Steroid Hormone Receptor Activity.

Previous studies demonstrated that the 52-kDa FK506-binding protein (FKBP52) proline-rich loop is functionally relevant in the regulation of steroid hormone receptor activity. While zebra fish (Danio rerio; Dr) FKBP52 contains all of the analogous domains and residues previously identified as critical for FKBP52 potentiation of receptor activity, it fails to potentiate activity. Thus, we used a cross-species comparative approach to assess the residues that are functionally critical for FKBP52 function. Random selection of gain-of-function DrFKBP52 mutants in Saccharomyces cerevisiae identified two critical residues, alanine 111 (A111) and threonine 157 (T157), for activation of receptor potentiation by DrFKBP52. In silico homology modeling suggests that alanine to valine substitution at position 111 in DrFKBP52 induces an open conformation of the proline-rich loop surface similar to that observed on human FKBP52, which may allow for sufficient surface area and increased hydrophobicity for interactions within the receptor-chaperone complex. A second mutation in the FKBP12-like domain 2 (FK2), threonine 157 to arginine (T157R), also enhanced potentiation, and the DrFKBP52-A111V/T157R double mutant potentiated receptor activity similar to human FKBP52. Collectively, these results confirm the functional importance of the FKBP52 proline-rich loop, suggest that an open conformation on the proline-rich loop surface is a predictor of activity, and highlight the importance of an additional residue within the FK2 domain.

Superheated liquid and supercritical denatured ethanol extraction of antioxidants from Crimson red grape stems.

Grapes are widely known for health benefits due to their antioxidant content. In wine production, grape stems are often discarded, though they has a higher content of antioxidants than the juice. The effectiveness of using an environmentally friendly solvent, ethanol, as a superheated liquid and supercritical fluid to extract antioxidant compounds from grape stems of organically grown Crimson Seedless grapes was evaluated. The Ferric Reducing Ability of Plasma (FRAP) assay and the Total Phenolic Content (TPC), or Folin-Ciocalteu assay, were used to quantify the antioxidant power of grape stem extracts. The extractions were performed at temperatures between 160°C and 300°C at constant density. It was found that the optimal extraction temperature was 204°C, at superheated liquid conditions, with a FRAP value of 0.670 mmol Trolox Equivalent/g of dry grape stem. The FRAP values were higher than other studies that extracted antioxidants from grape stems using single-pass batch extraction.