Proteinase location

Vos, P., van Asseldonka, M., van Jeveren, F., Siezen, R., Simons, G., and de Vos, W. M. (1989b). A maturation protein is essential for production of active forms of Lactococcus lactis SKI 1 serine proteinase located in or secreted from the cell envelope. J. Bacteriol. 171, 2795-2802. 

The expression of many of these molecules has been studied during various stages of differentiation of normal neutrophils and also of corresponding leukemic cells employing molecular biology techniques (eg, measurements of their specific mRNAs). For the majority, cDNAs have been isolated and sequenced, amino acid sequences deduced, genes have been localized to specific chromosomal locations, and exons and intron sequences have been defined. Some important proteinases of neutrophils are listed in Table 52-12. 

Based on their sequence homology, disulfide connectivity, and cysteine location within the sequence and chemistry of the reactive site. Pis can be assigned to distinct families, as classified by Laskowski and Kato. Kunitz-type, Bowman—Birk-type, Potato type I and type II, and squash inhibitors are members of these families shown in Table 3. For inhibitors not falling into these classifications more families have been proposed. Pis can also be classified by their target/mode of action. Plants have been found to express Pis that target serine proteinases, cysteine proteinases, aspartic proteinases, and metallo-proteinases. Serine and cysteine protease inhibitors are the best-studied PIs. ... 

Trypsin inhibitors in cucumber were first found in a study by Walker-Simmons et /. " after wounding of leaves and treatment with proteinase inhibitor-inducing factor (PIIF). The amino acid sequence of two inhibitors isolated from Cucurhita maxima (winter squash) were determined by Wilusz et at The peptides named ITD I and ITD 111 each comprised a 29-residue sequence with six cysteine residues. The only difference between the two peptides is in position 9, which is lysine in ITD I and glutamic acid in ITD III. The reactive site is located at the peptide bond between Arg5 and Ile6. Owing to their discovery and distribution in Cucurbitaceae the inhibitor family has been named squash inhibitors. Since the initial discoveries many other members of the squash family have been found. 

The functional proteins in the cell have to be protected in order to prevent premature degradation. Some of the intracellularly active proteolytic enzymes are therefore enclosed in lysosomes (see p. 234). The proteinases that act there are also known as cathepsins. Another carefully regulated system for protein degradation is located in the cytoplasm. This consists of large protein complexes (mass 2 10 Da), the proteasomes. Proteasomes contain a barrel-shaped core consisting of 28 subunits that has a sedimentation coef cient (see p. 200) of 20 S. Proteolytic activity (shown here by the scissors) is localized in the interior of the 20-S core and is therefore protected. The openings in the barrel are sealed by 19-S particles with a complex structure that control access to the core. 

Site 1 of proteinase K (2PRK) is located in the same region as site 2 of the subtilisins, but has different ligands again (Fig. 2g and h). It is one of the few octacoordinate protein Ca -binding sites observed to date, with... 

Lactic streptococci initiate casein degradation through the action of cell wall-associated and cell membrane-associated proteinases and peptidases. Small peptides are taken into the cell and hydrolyzed to their constituent amino acids by intracellular peptidases (Law and Sharpe 1978). Peptides containing four to seven residues can be transported into the cell by S. cremoris (Law et al. 1976B). S. lactis and S. cremoris have surface-bound peptidases and thus are not totally dependent on peptide uptake for protein use (Law 1979B). Some surface peptidases of S. cremoris are located in the cell membrane, whereas others are located at the cell wall-cell membrane interface (Exterkate 1984). Lactic streptococci have at least six different aminopeptidase activities, and can be divided into three groups based on their aminopeptidase profiles (Kaminogawa et al 1984). 

Argyls, P. J., Mathison, G. E. and Chandan, R. C. 1976. Production of cell-bound proteinase by Lactobacillus bulgaricus and its location in the bacteried cell. J. Appl. Bacte-riol 41, 175-184. 

Substrate is digested at sites where proteinases are located. 

A reaction looked at earlier simulates borate inhibition of serine proteinases.33 Resorufin acetate (234) is proposed as an attractive substrate to use with chymotrypsin since the absorbance of the product is several times more intense than that formed when the more usual p-nitrophcnyl acetate is used as a substrate. The steady-state values are the same for the two substrates, which is expected if the slow deacylation step involves a common intermediate. Experiments show that the acetate can bind to chymotrypsin other than at the active site.210 Brownian dynamics simulations of the encounter kinetics between the active site of an acetylcholinesterase and a charged substrate together with ah initio quantum chemical calculations using the 3-21G set to probe the transformation of the Michaelis complex into a covalently bound tetrahedral intermediate have been carried out.211 The Glu 199 residue located near the enzyme active triad boosts acetylcholinesterase activity by increasing the encounter rate due to the favourable modification of the electric field inside the enzyme and by stabilization of the TS for the first chemical step of catalysis.211... 

Fast-atom-bombardment mass spectrometry of a proteinase-truncated glucosyl-enzyme complex showed that the glucose was covalently linked as a /3-acetal ester to the carboxyl group of the aspartic acid located at the sixth residue from the N-terminal of the peptide ... 

We have shown above, although in a very qualitative way, that various NMR evidence seems to indicate that two peculiar features of SSI, namely dimeric structure and wide inhibitory activity, which are rather unusual for protein proteinase inhibitors, are actually closely related. The molecular assembly of the SSI subunit can be conceptually divided into two subdomains having a similar topology domain I, which is located at the N-... 

Aqualysin I contains two disulfide bonds which are formed between Cys67 and Cys99, and between Cysl63 and Cysl94 (Fig. 12.2).433 Proteinase K contains one free cysteine residue which is located at the corresponding position to the free cysteine residue of thermitase, and the other four cysteine residues form two disulfide bonds between two N-terminal-side residues and between two other C-terminal-side residues.443 Positions of the disulfide bond-forming cysteine residues of the two enzymes are different in the primary and... 

In our initial research on semisynthetic enzymes, we examined briefly the modification of the serine proteinase a-chymotrypsin, perhaps the best understood of the proteolytic enzymes. A logical choice as a residue for alkylation in the active site of a-chymotrypsin is His-57. However, an examination of a three-dimensional model (Lab Quip) of chymotrypsin in which coenzyme analogs were covalently attached to His-57 suggested strongly that such modifications would block completely the enzyme s active site region and that the probability of new reactions being catalyzed by the modified enzyme would be low. Another possible site of modification of chymotrypsin that could be considered was Met-192. This residue, located on the periphery of the... 

Internal water molecules may be located in the same sites in related proteins. In many proteins, internal waters are an essential part of the three-dimensional structure and their positions are typical for a whole family of proteins. Thus, in the three related plant cysteine proteinases, actinidin, papain, and calotropin Dl, 15 of the 16 internal waters (8 of which are illustrated in Fig. 19.13) are found in... 

Opaque cells produce the secreted aspartic proteinases Sap Ip, Sap2p and Sap3p in vitro, while the white form of the same strain cultured under identical conditions produces Sap2p only [22-24]. In an intravenous infection model, the white phenotype was more virulent than the opaque phenotype. In contrast, in a murine cutaneous candidiasis model, the opaque phenotype was more infectious [25]. In addition to yeast-hyphal dimorphism, phenotypic switching can provide a means for rapid adaptation of C. albicans to a changing environment in different anatomical locations. 

Hydrolysis of proteins without taste by proteases often produces bitter peptides. Hydrophobic amino acid residues located in the interior of protein molecules in aqueous solution are exposed by fragmentation of the protein molecules treated with proteases, and the peptides containing a number of hydrophobic amino acid residues occur in the solution (13). Many bitter peptides as shovm in Table 4-have been isolated from protein digests with proteinases (14-22). 

Fig. 6. Transmembrane arrangement of the a- and /3-polypeptides of the B880 antenna complex from Rs. rubrum. The hydrophobic domain is located within the hydrocarbon-tail region of the membrane the N- and C-terminal domains are at or near the membrane surface at the cytoplasmic or periplasmic side, respectively. PK, proteinase K CH, chymotrypsin TR, trypsin SA, 5. aureus protease.

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