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Protein Engineering Jan 1994The trypsin sequences currently available in the data banks have been collected and aligned using first the amino acid sequence homology and, subsequently, the... (Comparative Study)
Comparative Study
The trypsin sequences currently available in the data banks have been collected and aligned using first the amino acid sequence homology and, subsequently, the superposed crystal structures of trypsins from the cow, the bacterium Streptomyces griseus and the fungus Fusarium oxysporum. The phylogenetic tree constructed according to this multiple alignment is consistent with a continuous evolutionary divergence of trypsin from a common ancestor of both prokaryotes and eukaryotes. Comparison of crystal structures reveals a strict conservation of secondary structure. Similarly, in the alignment of all the sequences, insertions and deletions occur only in regions corresponding to loops between the secondary structure elements in the known crystal structures. The conserved residues cluster around the active site. Almost all conserved residues can be associated with one of the basic functional features of the protein: zymogen activation, catalysis and substrate specificity. In contrast, the residues of the hydrophobic core of the protein and the calcium ion binding sites are generally not conserved. The conserved features of trypsin and the nature of the conservation are discussed in detail.
Topics: Amino Acid Sequence; Biological Evolution; Consensus Sequence; Crystallography; Enzyme Activation; Molecular Sequence Data; Phylogeny; Protein Conformation; Protein Structure, Secondary; Sequence Alignment; Sequence Homology, Amino Acid; Species Specificity; Structure-Activity Relationship; Trypsin
PubMed: 8140095
DOI: 10.1093/protein/7.1.57 -
Protein Engineering Jun 1993The trypsin from Fusarium oxysporum is equally homologous to trypsins from Streptomyces griseus, Streptomyces erythraeus and to bovine trypsin. A DFP... (Comparative Study)
Comparative Study
The trypsin from Fusarium oxysporum is equally homologous to trypsins from Streptomyces griseus, Streptomyces erythraeus and to bovine trypsin. A DFP (diisopropylfluorophosphate) inhibited form of the enzyme has been crystallized from 1.4 M Na2SO4, buffered with citrate at pH 5.0-5.5. The crystals belong to space group P2(1) with cell parameters a = 33.43 A, b = 67.65 A, c = 39.85 A and beta = 107.6 degrees. There is one protein molecule in the asymmetric unit. X-ray diffraction data to a resolution of 1.8 A were collected on film using synchrotron radiation. The structure was solved by molecular replacement using models of bovine and S. griseus trypsins and refined to an R-factor of 0.141. The overall fold is similar to other trypsins, with some insertions and deletions. There is no evidence of the divalent cation binding sites seen in other trypsins. The covalently bound inhibitor molecule is clearly visible.
Topics: Amino Acid Sequence; Animals; Aspergillus oryzae; Base Sequence; Binding Sites; Cattle; Crystallization; Fusarium; Models, Molecular; Molecular Sequence Data; Molecular Structure; Recombinant Proteins; Streptococcus; Transformation, Bacterial; Trypsin; X-Ray Diffraction
PubMed: 8332590
DOI: 10.1093/protein/6.4.341 -
Biochimica Et Biophysica Acta Feb 1960
Topics: Acetates; Acetylation; Protein Processing, Post-Translational; Trypsin
PubMed: 13845801
DOI: 10.1016/0006-3002(60)91197-5 -
Marine Biotechnology (New York, N.Y.) 2005Atlantic cod trypsin I is an appropriate representative of the traditionally classified cold-adapted group I trypsins, and the recombinant form of cod trypsin Y is the... (Comparative Study)
Comparative Study Review
Atlantic cod trypsin I is an appropriate representative of the traditionally classified cold-adapted group I trypsins, and the recombinant form of cod trypsin Y is the only biochemically characterized member of the novel group III trypsins. Trypsin Y is adapted to lower temperatures than all other presently known trypsins. This review describes the basic characteristics of and practical uses for trypsins of Atlantic cod, as well as those of other organisms. Overexpression of the recombinant forms of cod trypsins I and Y in microorganisms is explained as well as the advantages of using site-directed mutagenesis to increase their stability toward autolysis and thermal inactivation. Trypsins appear to play a key role in the nutrition and development of marine fish. We discuss the potential use of cod trypsins as biomarkers to evaluate the nutritional status of cod larvae and describe the industrial applications of cod trypsin I and other trypsins.
Topics: Adaptation, Physiological; Amino Acid Sequence; Animals; Biotechnology; Cold Temperature; Escherichia coli; Fisheries; Fishes; Gene Expression; Iceland; Models, Molecular; Molecular Sequence Data; Mutagenesis, Site-Directed; Pichia; Sequence Alignment; Trypsin
PubMed: 15759084
DOI: 10.1007/s10126-004-0061-9 -
The Journal of Biological Chemistry Apr 1960
Topics: Catalysis; Chymotrypsin; Trypsin
PubMed: 13852782
DOI: No ID Found -
Journal of Agricultural and Food... May 2007Trypsin from the pyloric ceca of Atlantic bonito (Sarda sarda) was purified and characterized with respect to its purity; molecular weight; sensitivity to temperature,...
Trypsin from the pyloric ceca of Atlantic bonito (Sarda sarda) was purified and characterized with respect to its purity; molecular weight; sensitivity to temperature, pH, and inhibition; and N-terminal sequence. The purified trypsin had a molecular weight of 29 kDa as per sodium dodecyl sulfate polyacrylamide gel electrophoresis, and optimal activity was observed at pH 9 and 65 degrees C with BAPNA as a substrate. The enzyme was stable to heat treatment up to 50 degrees C and within the pH range of 7-12. It was stabilized by calcium ions, but its activity was strongly inhibited by soybean trypsin inhibitor, N-p-tosyl-L-lysine chloromethyl ketone, and phenyl methyl sulfonyl fluoride. The enzyme exhibited a progressive decrease in activity with increasing NaCl concentration (0-30%). The N-terminal 20 amino acid residues of Atlantic bonito trypsin were determined as IVGGYECQAHSQPWQPVLNS and were homologous with other trypsins.
Topics: Amino Acid Sequence; Animals; Cecum; Enzyme Stability; Molecular Sequence Data; Molecular Weight; Perciformes; Sequence Alignment; Sequence Analysis, Protein; Trypsin; Trypsin Inhibitors
PubMed: 17469841
DOI: 10.1021/jf063319x -
Biomacromolecules Dec 2010Polymer conjugation increases an enzyme's circulation time and stability for use as a therapeutic agent, but this attachment indubitably affects its properties. Covalent...
Polymer conjugation increases an enzyme's circulation time and stability for use as a therapeutic agent, but this attachment indubitably affects its properties. Covalent attachment of multiple polyethylene glycol chains with sizes of either 2, 5, 10, or 20 kDa increases the molecular weight and hydrodynamic radius of the model enzyme trypsin. The sizes of these polymer-enzyme conjugates are increased to be within the recommended limits for PDEPT applications. The T(d) increases from 49 to 60 °C to expand the enzyme's workable range of conditions. This functionalization with PEG polymers of varying lengths maintains trypsin's enzymatic activity. Conjugate activities are 79-120% that of native trypsin at room temperature and 221-432% that of trypsin at 37 °C.
Topics: Enzyme Stability; Molecular Weight; Particle Size; Polyethylene Glycols; Polymers; Trypsin
PubMed: 20979350
DOI: 10.1021/bm1006954 -
Experimental Cell Research Aug 1959
Topics: Tissue Culture Techniques; Trypsin
PubMed: 14405274
DOI: 10.1016/0014-4827(59)90300-3 -
Comparative Biochemistry and... 19881. Two trypsin-like enzymes, designated Trypsin A and B, were purified from the pyloric caeca and intestine of anchovy by (NH4)2SO4 fractionation, affinity...
1. Two trypsin-like enzymes, designated Trypsin A and B, were purified from the pyloric caeca and intestine of anchovy by (NH4)2SO4 fractionation, affinity chromatography (Benzamidine-Sepharose-6B) and ion exchange chromatography (DEAE-Sepharose). 2. Both trypsins catalyzed the hydrolysis of N-benzoyl-DL-arginine p-nitroanilide (BAPNA), p-tosyl-L-arginine methyl ester (TAME), casein and myofibrillar protein and they were inhibited by several well established trypsin-inhibitors. 3. The enzymes had mol. wts of 27,000 (Trypsin A) and 28,000 (Trypsin B). Their isoelectric points were about 4.9 (Trypsin A) and 4.6 (Trypsin B) and they had similar amino acid composition. 4. The enzymes had a pH optimum of 8-9 for the hydrolysis of BAPNA and of 9.5 for the digestion of casein and myofibrillar protein. Their activity and stability were affected by calcium ions. 5. Trypsins A and B resemble other fish trypsins in their mol. wt, pI, kinetic properties and the instability at low pH and they are similar to bovine trypsin in their dependence of Ca2+ for activity and stability.
Topics: Amino Acids; Animals; Digestive System; Fishes; Hydrogen-Ion Concentration; In Vitro Techniques; Isoelectric Point; Kinetics; Molecular Weight; Substrate Specificity; Temperature; Trypsin; Trypsin Inhibitors
PubMed: 3224506
DOI: 10.1016/0305-0491(88)90191-5 -
Biochimica Et Biophysica Acta Feb 1960
Topics: Arginine; Dioxanes; Hydrolysis; Kinetics; Trypsin; Water
PubMed: 13852783
DOI: 10.1016/0006-3002(60)91196-3