Trypsin peptides

Triterpenoid, 1071 tRNA, see Transfer RNA Trypsin, peptide cleavage with, 1033 Tryptophan, pKa of, 52... 

FIGURE 2-9. This peptide profile represents 16 hours of tryptic digestion. Note that peak 6 has decreased in height while 2 and 4 have increased. The peptide corresponding to peak 6 contains an internal lysine (K) which is slowly cleaved by trypsin. Peptide 8 also contains an internal lysine (K) which is well-protected from cleavage. As a result peaks X and Y (cleavage products of peptide 8) are only produced after long exposure to trypsin. (Reprinted from reference 6 with permission.)... 

Figure 1 5 MALDI spectrum from a 2-D gel spot excised from a human proteomic study in which the corresponding spectrum of the cathepsin D precursor could be identified after using SMEC micropreparation sample preparation followed by elution and spotting onto the MALDI target plate and MALDI analysis. The peptide mass fingerprinting revealed the identity of the protein using the Mascot bioinformatic software and the Swissprot protein database. The ( ) indicates the peptide masses corresponding to the cathepsin D precursor, and (T) the trypsin peptide fragments that were used for internal mass calibration.

Excised (2) Trypsin Peptide proteins digestion mixture... 

Trypsin peptides, keratin peptides (human or animal) siloxanes and PEG... 

After enzymatic digestion (trypsin), peptides are analyzed by LC-MS/MS... 

With /J-glucosidase As from A. wentii, difficulties arise from the tendency of the large cyanogen bromide and trypsin peptides to form insoluble aggregates. These could be overcome by carrying out the first chromatographic separation in 10% acetic acid on Sephadex G-50. 

The proteinases chymotrypsin and trypsin are two enzymes for which secondary and tertiary structures have been elucidated by x-ray analysis and which have structures supporting the lock and key hypothesis to a certain extent. The binding site in chymotrypsin and trypsin is a three-dimensional hydrophobic pocket (Fig. 2.11). Bulky amino acid residues such as aromatic amino acids fit neatly into the pocket (chymotrypsin. Fig. 2.11a), as do substrates with lysyl or arginyl residues (trypsin. Fig. 2.11b). Instead of Ser, the trypsin peptide chain has Asp which is present in the deep cleft in the form of a carboxylate anion and which attracts the positively charged lysyl or arginyl residues of the substrate. Thus, the substrate is stabilized and realigned by its peptide bond to face the enzyme s Ser which participates in hydrolysis (transforming locus). 

Fig. 5. Representative peptide mass fingerprint for the 33.2 and 66.6 kDa proteins resulting from an in-gel digestion of the 66.6 kDa gel band obtained by MALDI-TOF mass spectrometry. Peptides marked with an asterisk result from autodigestion products of recombinant trypsin. Peptides labeled with a diamond were selected for MS/MS experiments carried out on a MALDI-QIT/RTOF and a nano-ESI-QIT mass spectrometer.

The peptide shown m green was isolated by trypsin catalyzed hydrolysis and has an ammo acid sequence that completes the remaining overlaps... 

Trypsin (Section 27 10) A digestive enzyme that catalyzes the hydrolysis of proteins Trypsin selectively catalyzes the cleavage of the peptide bond between the carboxyl group of lysine or arginine and some other amino acid... 

Fig. 1. Schematic drawing of precursors for selected brain oligopeptides. Shaded areas represent the location of sequences of active peptide products which are normally cleaved by trypsin-like enzymes acting on double-basic amino acid residues. Precursors are not necessarily drawn to scale, (a) CRF precursor (b) proopiomelanocortin (POMC) (c) P-protachykinin (d) proenkephalin A (e) CGRP precursor (f) preprodynorphin, ie, preproenkephalin B. Terms are...

Group II consists of the enkephalins which come from the 267-aniino acid piecuisoi pro-enkephalin A [88402-54-4] (Fig. 2). This proteia contains four copies of Met-enkephalin, one copy of Leu-enkephalin, and the extended peptides Met-enkephalin-Arg -Phe (the last Met-enkephalin sequence ia Fig. 2) and Met-enkephalin-Arg -Gly -Leu (the fourth Met-enkephalin sequence ia Fig. 2) (25,26). AH of these products ate formed by trypsin-like cleavage between pairs of basic residues. The extended enkephalin peptides are further cleaved by carboxypeptidase E (27) to form authentic Met-enkephalin. 

Even though these enzymes have no absolute specificity, many of them show a preference for a particular side chain before the scissile bond as seen from the amino end of the polypeptide chain. The preference of chymotrypsin to cleave after large aromatic side chains and of trypsin to cleave after Lys or Arg side chains is exploited when these enzymes are used to produce peptides suitable for amino acid sequence determination and fingerprinting. In each case, the preferred side chain is oriented so as to fit into a pocket of the enzyme called the specificity pocket. 

FIGURE 5.20 Trypsin is a proteolytic enzyme, or protease, that specifically cleaves only those peptide bonds in which arginine or lysine contributes the carbonyl function. The products of the reaction are a mixture of peptide fragments with C-terminal Arg or Lys residues and a single peptide derived from the polypeptide s C-terminal end. 

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