Oxidative Deamination Activity of EGCG

(-)-Epigallocatechin-3-O-gallate (EGCG), the most abundant polyphenol in green tea, mediates the oxidative modification of proteins, generating protein carbonyls. However, the underlying molecular mechanism remains unclear. Here we analyzed the EGCG-derived intermediates generated upon incubation with the human serum albumin (HSA) and established that EGCG selectively oxidized the lysine residues via its oxidative deamination activity. In addition, we characterized the EGCG-oxidized proteins and discovered that the EGCG could be an endogenous source of the electrically-transformed proteins that could be recognized by the natural antibodies. When HSA was incubated with EGCG in the phosphate-buffered saline (pH 7.4) at 37°C, the protein carbonylation was associated with the formation of EGCG-derived products, such as the protein-bound EGCG, oxidized EGCG, and aminated EGCG. The aminated EGCG was also detected in the sera from the mice treated with EGCG in vivo. EGCG selectively oxidized lysine residues at the EGCG-binding domains in HSA to generate an oxidatively deaminated product, aminoadipic semialdehyde. In addition, EGCG treatment results in the increased negative charge of the protein due to the oxidative deamination of the lysine residues. More strikingly, the formation of protein carbonyls by EGCG markedly increased its cross-reactivity with the natural IgM antibodies. These findings suggest that many of the beneficial effects of EGCG may be partly attributed to its oxidative deamination activity, generating the oxidized proteins as a target of natural antibodies

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

The Mediterranean diet: Effects on proteins that mediate fatty acid metabolism in the colon

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/88101/1/j.1753-4887.2011.00439.x.pd

Mimicking Wagyu beef fat in cultured meat: Progress in edible bovine adipose tissue production with controllable fatty acid composition

Since the current process of livestock meat production has considerable effects on the global environment, leading to high emissions of greenhouse gases, cultured meat has recently attracted attention as a suitable alternative way to acquire animal proteins. However, while most published studies on cell-cultured meat have focused on muscle tissue culture, fat production which is an important component of the process has often been neglected from this technology, even though it can enhance the meat's final taste, aroma, tenderness, texture, and palatability. In this study, we focused on bovine muscle reconstruction by monitoring and optimizing the possible expansion rate of isolated primary bovine adipose stem cells and their adipogenesis differentiation to be fully edible for cultured meat application.After approximatively 100 days of serial passages, the bovine adipose-derived stem cells, isolated from muscle tissue, underwent 57 ​± ​5 doublings in the edible cell culture medium condition. This implies that by cultivating and amplifying them, a minimum of 2.9 ​× ​1022 ​cells can be obtained from around 10 ​g of fat. It was discovered that these cells retain their adipogenesis differentiation ability for at least 12 passages. Moreover, the final lipid composition could be controlled by adjusting the fatty acid composition of the culture medium during the differentiation process, resulting in organoleptic features similar to those of real fat from muscle. This was especially so for the cis isomer oleic acid percentage, an important part of high-grade Japanese Wagyu meat.These characteristics of the primary bovine adipose-derived stem cell proliferation and adipogenesis differentiation provide valuable insights for the in vitro production of meat alternatives

Identification of Polyphenol-Specific Innate Epitopes That Originated from a Resveratrol Analogue

Polyphenols have received a significant amount of attention in disease prevention because of their unique chemical and biological properties. However, the underlying molecular mechanism for their beneficial effects remains unclear. We have now identified a polyphenol as a source of innate epitopes detected in natural IgM and established a unique gain-of-function mechanism in the formation of innate epitopes by polyphenol via the polymerization of proteins. Upon incubation with bovine serum albumin (BSA) under physiological conditions, several polyphenols converted the protein into the innate epitopes recognized by the IgM Abs. Interestingly, piceatannol, a naturally occurring hydroxylated analogue of a red wine polyphenol, resveratrol, mediated the modification of BSA, whose polymerized form was specifically recognized by the IgMs. The piceatannol-mediated polymerization of the protein was associated with the formation of a lysine-derived cross-link, dehydrolysinonorleucine. In addition, an oxidatively deaminated product, α-aminoadipic semialdehyde, was detected as a potential precursor for the cross-link in the piceatannol-treated BSA, suggesting that the polymerization of the protein might be mediated by the oxidation of a lysine residue by piceatannol followed by a Schiff base reaction with the ε-amino group of an unoxidized lysine residue. The results of this study established a novel mechanism for the formation of innate epitopes by small dietary molecules and support the notion that many of the beneficial effects of polyphenols could be attributed, at least in part, to their lysyl oxidase-like activity. They also suggest that resveratrol may have beneficial effects on human health because of its conversion to piceatannol

LC-ESI-MS/MS analysis of oxidized amino acids in the EGCG-treated HSA.

<p>(<b>A</b>) Chemical structures of aminoadipic semialdehyde (AAS) and glutamic semialdehyde (GGS). (<b>B</b>) Collision-induced dissociation of the [M+H]⁺ of ABA-AAS at <i>m/z</i> 267 at a collision energy of 25 V and the proposed structures of individual ions. (<b>C</b>) Collision-induced dissociation of the [M+H]⁺ of ABA-GGS at <i>m/z</i> 253 at a collision energy of 25 V and the proposed structures of individual ions. (<b>D</b>) The ion current tracings of ABA-AAS (<i>left three tracings</i>) and ABA-GGS (<i>right three tracings</i>) using LC-ESI-MS/MS with SRM. (<b>E</b>) Determination of AAS in the EGCG-treated HSA. HSA (1 mg/ml) was incubated with EGCG (1 mM) in 0.1 ml of PBS (pH 7.4) for 24 h at 37°C. The yield of AAS was semi-quantitatively determined based on a calibration curve (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0153002#pone.0153002.s003" target="_blank">S3 Fig</a>). (<b>F</b>) Schematic illustration of the transformation of a lysine residue to AAS by EGCG.</p

LC-ESI-MS/MS analysis of EGCG and NH₂-EGCG in the sera of mice treated with EGCG.

<p>(<b>A</b>) The ion current tracings of EGCG (<i>left five tracings</i>) and NH₂-EGCG (<i>right five tracings</i>). BALB/c mice were intraperitoneally injected with 0.1 ml of EGCG (10 mM) or PBS. After injection for 10 or 30 min, the sera were collected. After removing proteins by precipitation with cold acetone, the samples were analyzed by LC-ESI-MS/MS with SRM mode. (<b>B</b>) Quantitative analysis of EGCG (<i>upper panel</i>) and NH₂-EGCG (<i>lower panel</i>) in the sera of the mice treated with EGCG.</p

Oxidative Deamination of Serum Albumins by (-)-Epigallocatechin-3-<i>O</i>-Gallate: A Potential Mechanism for the Formation of Innate Antigens by Antioxidants

<div><p>(-)-Epigallocatechin-3-<i>O</i>-gallate (EGCG), the most abundant polyphenol in green tea, mediates the oxidative modification of proteins, generating protein carbonyls. However, the underlying molecular mechanism remains unclear. Here we analyzed the EGCG-derived intermediates generated upon incubation with the human serum albumin (HSA) and established that EGCG selectively oxidized the lysine residues via its oxidative deamination activity. In addition, we characterized the EGCG-oxidized proteins and discovered that the EGCG could be an endogenous source of the electrically-transformed proteins that could be recognized by the natural antibodies. When HSA was incubated with EGCG in the phosphate-buffered saline (pH 7.4) at 37°C, the protein carbonylation was associated with the formation of EGCG-derived products, such as the protein-bound EGCG, oxidized EGCG, and aminated EGCG. The aminated EGCG was also detected in the sera from the mice treated with EGCG <i>in vivo</i>. EGCG selectively oxidized lysine residues at the EGCG-binding domains in HSA to generate an oxidatively deaminated product, aminoadipic semialdehyde. In addition, EGCG treatment results in the increased negative charge of the protein due to the oxidative deamination of the lysine residues. More strikingly, the formation of protein carbonyls by EGCG markedly increased its cross-reactivity with the natural IgM antibodies. These findings suggest that many of the beneficial effects of EGCG may be partly attributed to its oxidative deamination activity, generating the oxidized proteins as a target of natural antibodies.</p></div

Docking simulations of EGCG to HSA.

<p>(<b>A</b>) Surface representation of HSA with the lysine residues sensitive to oxidation highlighted in yellow. (<b>B</b>) <i>Left</i>, schematic representation of the EGCG binding site in subdomains IIA and IIIA. Ribbon model is colored in orange. <i>Right</i>, close up view of the EGCG-binding pocket from its entrance located in subdomains IIA and IIIA. The electrostatic potential is represented on a color scale from blue for a positive potential, white for neutral, to red for a negative potential. (<b>C</b>) <i>Left</i>, schematic representation of the predicted EGCG binding site in subdomain IIIB. Ribbon model is colored in orange. <i>Right</i>, close up view of the predicted EGCG-binding pocket in subdomain IIIB. In panels <b>B</b> and <b>C</b>, selected residues shown in stick and color-coded by atom type: carbon in <i>dark green</i>; oxygen in <i>red</i>; and nitrogen in <i>blue</i>. The electrostatic potential is represented on a color scale from blue for a positive potential, white for neutral, to red for a negative potential. The EGCG molecule is shown in stick and color-coded by atom type: carbon in <i>right green</i>; oxygen in <i>red</i>.</p

Formation of electrically-charged proteins by EGCG.

<p>(<b>A</b>) Changes in the zeta potential of HSA treated with the catechins. HSA (1 mg/ml) was incubated with 1 mM catechins in 0.1 ml of PBS (pH 7.4) for 24 h at 37°C. (<b>B</b>) Changes in the zeta potential of BSA treated with the metal-catalyzed oxidation reactions. BSA (1 mg/ml) was incubated with 200 μM PQQ or 1 mM H₂O₂ in the presence and absence of 100 μM Cu²⁺ in 0.1 ml of PBS (pH 7.4) for 24 h at 37°C. (<b>C</b>) Schematic illustration of the EGCG-mediated transformation of HSA into electronegative molecules via oxidative deamination.</p