Dataset Information

Pseudotrypsin: A Little-Known Trypsin Proteoform.

ABSTRACT: Trypsin is the protease of choice for protein sample digestion in proteomics. The most typical active forms are the single-chain ?-trypsin and the two-chain ?-trypsin, which is produced by a limited autolysis of ?-trypsin. An additional intra-chain split leads to pseudotrypsin (?-trypsin) with three chains interconnected by disulfide bonds, which can be isolated from the autolyzate by ion-exchange chromatography. Based on experimental data with artificial substrates, peptides, and protein standards, ?-trypsin shows altered kinetic properties, thermodynamic stability and cleavage site preference (and partly also cleavage specificity) compared to the above-mentioned proteoforms. In our laboratory, we have analyzed the performance of bovine ?-trypsin in the digestion of protein samples with a different complexity. It cleaves predominantly at the characteristic trypsin cleavage sites. However, in a comparison with common tryptic digestion, non-specific cleavages occur more frequently (mostly after the aromatic residues of Tyr and Phe) and more missed cleavages are generated. Because of the preferential cleavages after the basic residues and more developed side specificity, which is not expected to occur for the major trypsin forms (but may appear anyway because of their autolysis), ?-trypsin produces valuable information, which is complementary in part to data based on a strictly specific trypsin digestion and thus can be unnoticed following common proteomics protocols.

SUBMITTER: Perutka Z

PROVIDER: S-EPMC6222510 | biostudies-literature | 2018 Oct

REPOSITORIES: biostudies-literature

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Pseudotrypsin: A Little-Known Trypsin Proteoform.

Perutka Zdeněk Z Šebela Marek M

Molecules (Basel, Switzerland) 20181014 10

Trypsin is the protease of choice for protein sample digestion in proteomics. The most typical active forms are the single-chain β-trypsin and the two-chain α-trypsin, which is produced by a limited autolysis of β-trypsin. An additional intra-chain split leads to pseudotrypsin (ψ-trypsin) with three chains interconnected by disulfide bonds, which can be isolated from the autolyzate by ion-exchange chromatography. Based on experimental data with artificial substrates, peptides, and protein standa ...[more]

PMID: 30322187

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Project description:Reactive oxygen species (ROS) are constantly produced by metabolically active plant cells. The concentration of ROS may determine their role, e.g., they may participate in signal transduction or cause oxidative damage to various cellular components. To ensure cellular homeostasis and minimize the negative effects of excess ROS, plant cells have evolved a complex antioxidant system, which includes ascorbic acid (AsA). AsA is a multifunctional metabolite with strong reducing properties that allows the neutralization of ROS and the reduction of molecules oxidized by ROS in cooperation with glutathione in the Foyer-Halliwell-Asada cycle. Antioxidant enzymes involved in AsA oxidation and reduction switches evolved uniquely in plants. Most experiments concerning the role of AsA have been performed on herbaceous plants. In addition to extending our understanding of this role in additional taxa, fundamental knowledge of the complex life cycle stages of woody plants, including their development and response to environmental factors, will enhance their breeding and amend their protection. Thus, the role of AsA in woody plants compared to that in nonwoody plants is the focus of this paper. The role of AsA in woody plants has been studied for nearly 20 years. Studies have demonstrated that AsA is important for the growth and development of woody plants. Substantial changes in AsA levels, as well as reduction and oxidation switches, have been reported in various physiological processes and transitions described mainly in leaves, fruits, buds, and seeds. Evidently, AsA exhibits a dual role in the photoprotection of the photosynthetic apparatus in woody plants, which are the most important scavengers of ozone. AsA is associated with proper seed production and, thus, woody plant reproduction. Similarly, an important function of AsA is described under drought, salinity, temperature, light stress, and biotic stress. This report emphasizes the involvement of AsA in the ecological advantages, such as nutrition recycling due to leaf senescence, of trees and shrubs compared to nonwoody plants.

Dataset Information

Pseudotrypsin: A Little-Known Trypsin Proteoform.

Publications

Pseudotrypsin: A Little-Known Trypsin Proteoform.

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