Differential impact of cold and hot tea extracts on tyrosine phosphatases regulating insulin receptor activity: a focus on PTP1B and LMW-PTP.

PURPOSE: The impact of tea constituents on the insulin-signaling pathway as well as their antidiabetic activity are still debated questions. Previous studies suggested that some tea components act as Protein Tyrosine Phosphatase 1B (PTP1B) inhibitors. However, their nature and mechanism of action remain to be clarified. This study aims to evaluate the effects of both tea extracts and some of their constituents on two main negative regulators of the insulin-signaling pathway, Low-Molecular-Weight Protein Tyrosine Phosphatase (LMW-PTP) and PTP1B. METHODS: The effects of cold and hot tea extracts on the enzyme activity were evaluated through in vitro assays. Active components were identified using gas chromatography-mass spectrometry (GC-MS) analysis. Finally, the impact of both whole tea extracts and specific active tea components on the insulin-signaling pathway was evaluated in liver and muscle cells. RESULTS: We found that both cold and hot tea extracts inhibit LMW-PTP and PTP1B, even if with a different mechanism of action. We identified galloyl moiety-bearing catechins as the tea components responsible for this inhibition. Specifically, kinetic and docking analyses revealed that epigallocatechin gallate (EGCG) is a mixed-type non-competitive inhibitor of PTP1B, showing an IC50 value in the nanomolar range. Finally, in vitro assays confirmed that EGCG acts as an insulin-sensitizing agent and that the chronic treatment of liver cells with tea extracts results in an enhancement of the insulin receptor levels and insulin sensitivity. CONCLUSION: Altogether, our data suggest that tea components are able to regulate both protein levels and activation status of the insulin receptor by modulating the activity of PTP1B.

Analytic investigations on protein content in refined seed oils: implications in food allergy.

BACKGROUND: A number of scientific reports have investigated the possible implications of refined seed oils in allergic reactions, resulting in conflicting points of view. Also the total amount of residual proteins after refinement is still a matter of debate. Nevertheless, seed oils are now blamed as possible cause of allergic reactions. OBJECTIVE: To determine the true amount of proteins after oil refinement and to shed new lights on allergenic properties of refined seed oils. METHODS: We optimized a protein extraction procedure on several commercial refined seed oils. Both colorimetric and amino acid analysis were used to measure residual protein content. SDS-PAGE was also used for characterizations of protein band patterns. Sensitized child patients sera were tested by Western blot on PAGE-resolved proteins. RESULTS: Our extraction method proved to be effective and reproducible. Amino acid analysis resulted more accurate in determining the protein content with respect to colorimetric methods, indicating a higher protein content than that previously reported. IgE responsive residual proteins were found in peanut oil extracts. CONCLUSIONS: Our preliminary data suggest that fully refined seed oils should be taken into account in the context of allergic reactions and would benefit of further toxicological studies.

Cerato-platanin, the first member of a new fungal protein family: cloning, expression, and characterization.

The ascomycete Ceratocystis fimbriata, the causal agent of "canker stain disease," secretes a protein of 12.4 kDa that elicits phytoalexin synthesis and plant cell death. This protein, named cerato-platanin (CP), is also located in the cell walls of ascospores, hyphae, and conidia; it contains four cysteines (S-S bridged) and is moderately hydrophobic. The cp gene consists of a single exon and has 42 bp codifying for a signal peptide of 14 residues. The recombinant protein was obtained by cloning the cp gene of the mature protein in Escherichia coli (BL21), and a refolding step was needed to achieve the native active form. In the European Molecular Biology data bank, CP is reported as the first member of the CP family; this is the first example of an set of secreted fungal proteins whose primary structure is very similar. Nonetheless, the data also revealed some structural and functional features that make CP similar to proteins of the hydrophobin family.