Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Twenty-two nitrogen-fixing Bacillus azotofixans strains were shown to produce an inhibition zone against themselves in plate assays. The B. azotofixans type strain P3L-5, chosen for further studies, produced inhibition zones against various Bacillus strains and other bacterial genera. This antibacterial substance was also produced in liquid medium and its production was enhanced in semisolid medium (0.4% agar) after 3 to 5 days of incubation. The substance was suggested to be an antibiotic and its preliminary characterization showed resistance to heat (100 degrees C, 15 minutes), to trypsin, pronase, deoxyribonuclease I, ribonuclease A, phospholipase C, ethanol, acetone, and ether, and sensitivity to strong alkali treatment. Its molecular weight was estimated to be between 3500 to 6000. After induction of B. azotofixans P3L-5 with mitomycin C or ultraviolet light, two types of particles were detected in the lysate: one similar to a phage tail and the other, less frequent, similar to a complete bacteriophage. Lysates containing these particles showed a killing effect in some but not all B. azotofixans strains, but neither the other Bacillus species nor Micrococcus were inhibited by these lysates.
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PMID:Production of a bacteriophage, a phage tail-like bacteriocin and an antibiotic by Bacillus azotofixans. 212

Infrared spectra have been obtained for 12 globular proteins in aqueous solution at 20 degrees C. The proteins studied, which vary widely in the relative amounts of different secondary structures present, include myoglobin, hemoglobin, immunoglobulin G, concanavalin A, lysozyme, cytochrome c, alpha-chymotrypsin, trypsin, ribonuclease A, alcohol dehydrogenase, beta 2-microglobulin, and human class I major histocompatibility complex antigen A2. Criteria for evaluating how successfully the spectra due to liquid and gaseous water are subtracted from the observed spectrum in the amide I region were developed. Comparisons of second-derivative amide I spectra with available crystal structure data provide both qualitative and quantitative support for assignments of infrared bands to secondary structures. Band frequency assignments assigned to alpha-helix, beta-sheet, unordered, and turn structures are highly consistent among all proteins and agree closely with predictions from theory. alpha-Helix and unordered structures can each be assigned to only one band whereas multiple bands are associated with beta-sheets and turns. These findings demonstrate a method of analysis of second-derivative amide I spectra whereby the frequencies of bands due to different secondary structures can be obtained. Furthermore, the band intensities obtained provide a useful method for estimating the relative amounts of different structures.
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PMID:Protein secondary structures in water from second-derivative amide I infrared spectra. 215 34

T. brucei cytoplasmic calcium-dependent alkaline ribonuclease activity from DEAE-cellulose fractionation was separated into endoribonuclease and exoribonuclease activities by hydroxyapatite chromatography. T. brucei cytoplasmic extract markedly decreased the endoribonuclease activity, but slightly potentiated the activities of the exoribonuclease and bovine ribonuclease A. While the endoribonuclease was activated by trypsin, the exoribonuclease and bovine ribonuclease A were partially inactivated by trypsin. The endoribonuclease was activated by p-chloromercuribenzoate or N-ethylmaleimide; the exoribonuclease was not affected by these sulfhydryl group reagents. Free ribonuclease was separated from the latent endoribonuclease by 1 M NaCl-Sephadex G-100 gel filtration. The results demonstrate that T. brucei cytoplasm contains a latent endoribonuclease consisting of ribonuclease and inhibitor protein.
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PMID:Trypanosoma brucei: calcium-dependent endoribonuclease is associated with inhibitor protein. 222 4

This study extends our previous observation that surface mannose receptor expression by pure populations of CSF-1-dependent bone marrow-derived macrophages increases with time (Clohisy, D. R., Bar-Shavit, Z., Chappel, J. C., and Teitelbaum, S. L. (1987) J. Biol. Chem. 262, 15922-15929). We presently find, however, that the progressive enhancement of 125I-mannose-bovine serum albumin (125I-Man-BSA) binding per cell reflects cell number rather than duration of culture. In fact, macrophages plated at high density bind 8-fold more 125I-Man-BSA than do their low density counterparts, with no difference in receptor-ligand affinity. Furthermore, cells cultured at high density are ultimately subjected to lower levels of exogenously provided macrophage growth factor, and fewer are in interphase. By obtaining synchronous populations of quiescent bone marrow macrophages, however, we demonstrate that neither cell cycling nor attendant levels of colony stimulating factor-1 influence mannose receptor expression. Our next series of experiments established that density-related mannose receptor expression reflects removal, by marrow macrophages, of a "down-regulating" factor contained in culture medium. To this end, we treated mononuclear phagocytes with either macrophage- or control-conditioned medium and found that, via a fetal calf serum-residing protein(s), only control medium is capable of noncompetitively reducing 125I-Man-BSA binding in a dose-dependent manner. Moreover, reconstituted 20-40% (NH4)2SO4-precipitable fractions derived from either sham-conditioned medium or fetal calf serum are capable of down-regulating mannose receptor expression. Alternatively, the same fraction obtained from macrophage-conditioned medium contains no such activity. Finally, initial characterization of the down-regulating factor reveals it to be acid-activable and trypsin-sensitive, yet resistant to heating to at least 80 degrees C, ribonuclease A, or freezing and thawing. We conclude that bone marrow macrophages up-regulate expression of their own plasma membrane mannose receptor by inactivating a noncompetitive, serum-residing inhibitory protein(s).
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PMID:Bone marrow-derived mononuclear phagocytes autoregulate mannose receptor expression. 292 10

The nature of vascular permeability factor (VPF) activity derived from serum-free conditioned medium containing cultured human malignant glial tumors has been further investigated. A 1000-fold purification was accomplished by sequential heparin-Sepharose affinity chromatography and high-performance liquid chromatography gel filtration chromatography steps. Vascular permeability factor activity falls into a molecular weight range of 41,000 to 56,000 D. Activity is bound to hydroxylapatite, carboxymethyl-Sepharose, phenyl-Sepharose, and heparin-Sepharose, whereas little or no activity was bound to diethylaminoethyl-Sephacel. Vascular permeability factor activity is trypsin- and pepsin-sensitive but is unaffected by treatment with ribonuclease A. This suggests that VPF is a hydrophobic, positively charged (cationic) polypeptide with a potentially biologically significant affinity for heparin. As most proteins are negatively charged (anionic) and have no affinity for heparin, a significant advantage was gained by performing these purification steps. The activity of VPF is not inhibited by coinjection of conditioned medium with soybean trypsin inhibitor; or hexadimethrine (both known antagonists of tissue plasminogen activator, Hageman factor, and serum kallikrein); or aprotinin (an antagonist of both plasmin and tissue kallikrein); or phenylmethanesulfonyl fluoride (a serine esterase (elastase) inhibitor); or pepstatin-A (an acid protease inhibitor which inactivates vascular permeability-inducing leukokinins). These data, together with the fact that VPF is produced and released into serum-free media, provides substantial evidence against it being one of the more commonly known serum-derived permeability mediators. Treatment with dithiothreitol inhibited VPF activity, indicating the presence of at least one essential disulfide bond in this molecule. Inhibition by dexamethasone of VPF expression in cultured malignant glial cells appears to be selective. Dexamethasone-induced inhibition of VPF was dose-responsive and was not associated with a parallel inhibition of cellular protein synthesis as determined by tritiated leucine incorporation into trichloroacetic acid-precipitable material. Inclusion of dexamethasone in the culture medium was not associated with altered cell viability or cell number. A series of in vivo studies confirmed the inhibition of VPF activity in test animals pretreated with dexamethasone. This steroid-induced inhibition was partially reversed by treatment of test animals with actinomycin D prior to exposure to dexamethasone.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Further characterization of malignant glioma-derived vascular permeability factor. 313 21

The primary structure of angiogenin is 33% identical to that of bovine pancreatic ribonuclease (RNase), but the enzymatic activities of the two proteins differ markedly. Similarly, their susceptibilities to limited proteolysis differ as well. In contrast to RNase, angiogenin totally resists proteolysis by subtilisin. Indeed, among 16 proteases examined, only endoprotease Lys-C, trypsin, and pepsin are able to cleave angiogenin. Even with prolonged incubation, endoprotease Lys-C selectively cleaves the Lys-60-Asn-61 bond; the product retains full ribonucleolytic activity. Initially, trypsin also cleaves this same bond, but with time it causes extensive degradation. Pepsin, at pH 2, cleaves the Phe-9-Leu-10 bond, to give angiogenin (10-123), which displays approximately 15% of the native activity toward ribosomal RNA (rRNA). The susceptibility to proteolysis and/or the sites of cleavage of angiogenin and bovine RNase differ markedly despite their structural homology. These differences are considered in terms of the amino acid sequences of the two proteins.
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PMID:Conformational characterization of human angiogenin by limited proteolysis. 315 Dec 51

A method for the prediction of hydrogen positions in proteins is presented. The method is based on the knowledge of the heavy atom positions obtained, for instance, from X-ray crystallography. It employs an energy minimization limited to the environment of the hydrogen atoms bound to a common heavy atom or to a single water molecule. The method is not restricted to proteins and can be applied without modification to nonpolar hydrogens and to nucleic acids. The method has been applied to the neutron diffraction structures of trypsin, ribonuclease A, and bovine pancreatic trypsin inhibitor. A comparison of the constructed and the observed hydrogen positions shows few deviations except in situations in which several energetically similar conformations are possible. Analysis of the potential energy of rotation of Lys amino and Ser, Thr, Tyr hydroxyl groups reveals that the conformations of lowest intrinsic torsion energies are statistically favored in both the crystal and the constructed structures.
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PMID:Polar hydrogen positions in proteins: empirical energy placement and neutron diffraction comparison. 322 15

The complete amino-acid sequence of BS-RNAse, a dimeric ribonuclease isolated from bovine seminal plasma, was determined. The reduced and S-carboxymethylated subunit chain of the enzyme was cleaved by trypsin and chymotrypsin. The resulting peptides, purified by cation-exchange chromatography were sequenced by dansyl-Edman, subtractive Edman degradation and carboxypeptidase A and B digestion. Chymotryptic peptides were used for the alignment. Automated Edman degradation of the native protein, through the N-terminal 41 amino-acid residues, completed the sequence information. The subunit chain of BS-RNAse, composed of 124 amino-acid residues, with a molecular mass of 13,610 Da, is highly homologous (81%) to pancreatic ribonuclease A. A good degree of homology (31%) was also found with human angiogenin. No N-linked carbohydrate-attachment sites, such as Asn-X-Ser/Thr, were found in the protein.
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PMID:Complete amino-acid sequence of bovine seminal ribonuclease, a dimeric protein from seminal plasma. 342 1

The mode of derivatization of amino groups of proteins by glyceraldehyde, an aldotriose, depends on the presence or absence of reducing agent. In the presence of sodium cyanoborohydride, the Schiff base adducts of the aldehyde with the amino groups are reduced, and dihydroxypropylation of amino groups takes place (reductive mode). The reductively glycated lysine residue, N epsilon-(2,3-dihydroxypropyl)lysine, is a substituted alpha-amino alcohol. This alpha-amino alcoholic function of the derivatized lysine should be susceptible to periodate oxidation, and this oxidation is anticipated to result in the regeneration of the lysine residue. This aspect has been now investigated. Indeed, on mild periodate oxidation (15 mM periodate, 15 min at room temperature) of dihydroxypropylated ribonuclease A, nearly 95% of its N epsilon-(2,3-dihydroxypropyl)lysine residues were regenerated to lysine residues. The removal of the dihydroxypropyl groups by periodate oxidation could be accomplished within a wide pH range with little variation in the recovery of lysines. The possible usefulness of this reversible chemical modification procedure in the primary structural studies of proteins was investigated with a tryptic peptide of dihydroxypropylated streptococcal M5 protein, namely, DHP-T4. This 12-residue tryptic peptide contains one internal N epsilon-(dihydroxypropyl)lysine. The dihydroxypropylated peptide released most of its dihydroxypropyl groups on mild periodate oxidation. Redigestion of the periodate-treated peptide with trypsin generated the two expected peptides, demonstrating the generation of a trypsin-susceptible site. Reductive dihydroxypropylation of amino groups of RNase A resulted in the loss of its enzyme activity, the extent of inactivation increasing with the concentration of the glyceraldehyde used.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Dihydroxypropylation of amino groups of proteins: use of glyceraldehyde as a reversible agent for reductive alkylation. 365 95

The chemical modification of bovine pancreatic ribonuclease A by 6-chloropurine riboside was studied to obtain information about the role of the purine nucleoside moiety of the ribonucleic acid in the enzyme-substrate interaction. The residues involved in the reaction were identified, after performic acid oxidation and trypsin digestion, by reverse-phase HPLC peptide mapping. The labeled peptides were detected by following the absorbance at 254 nm, and amino acid analyses of these peptides showed that the reaction had taken place with the amino groups of Lys-1, -37, -41, and -91. The specificity of the reaction was unaffected by changing the ligand:protein molar ratio. Partial separation of the reaction products was accomplished by means of chromatography on CM-Sepharose: four labeled fractions corresponding to mono- and bisubstituted derivatives were found. One of the monosubstituted fractions (fraction E) contained a homogeneous protein with the nucleoside bound to the alpha-amino group of Lys-1 whereas the other (fraction D) was a mixture of derivatives labeled in the epsilon-amino group of Lys-1, -37, -41, and -91. Kinetic studies of these two monosubstituted fractions were performed with cytidine 2',3'-phosphate and ribonucleic acid as substrates. These derivatives showed a noncompetitive inhibition-like behavior with respect to RNase A. Results support the existence of several RNase A regions with affinity for purine nucleosides.
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PMID:Modification of bovine pancreatic ribonuclease A with 6-chloropurine riboside. 370 27


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