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Y Birk

Publications and source records attributed to Y Birk.

At least 37 records · Page 2Linked to original sources
PubMed1984

Photoreactive derivative of Kunitz's soybean trypsin inhibitor. Preparation by selective modification of a tryptophan residue and formation of a covalent complex of the modified inhibitor with trypsin.

The photoreactive arylsulfenyl chloride 2-nitro-4-azidophenylsulfenyl chloride (2,4-NAPS-Cl) has been used for the selective modification of tryptophan in Kunitz's soybean trypsin inhibitor (SBTI). The ultraviolet absorption spectrum and amino acid analysis of 2,4-NAPS-SBTI indicated that only one of the two tryptophans (93 or 117) present in SBTI was modified. CNBr cleavage of 2,4-NAPS-SBTI resulted in two fragments 1-114 and 115-181. Amino acid analysis of the two separated fragments showed that only tryptophan 93 underwent modification. 2,4-NAPS-SBTI fully retained its inhibitory activity against trypsin. The photoaffinity labeling of trypsin with 2,4-NAPS-Cl was performed on tritiated trypsin prepared by reacting bovine trypsin with [3H]-succinimidyl propionate. The covalent attachment of 2,4-NAPS-SBTI to the tritiated trypsin after photolysis was demonstrated by exclusion chromatography on Sephadex G-50 in the presence of guanidine hydrochloride.

Affinity Labels↗
PubMed1983

Purification and properties of protease F, a bacterial enzyme with chymotrypsin and elastase specificities.

It has been previously demonstrated that commercial bacterial fibrinolysin (EC 3.4.21.7) selectively cleaves the bond between Met-53 and Ala-54 in ovine prolactin (199 amino acids). A one-step purification procedure on DEAE-cellulose for Protease F, which is the active component of bacterial fibrinolysin, and properties of the purified enzyme are reported. The enzyme is homogeneous as judged by acrylamide gel electrophoresis. Its molecular weight, calculated from gel filtration experiments on Sephadex G-100, is around 13,800. Amino acid analyses do not reveal the presence of any half-cystines. The presence of one tryptophan residue per enzyme molecule was resolved from the fluorescence spectrum. Amino terminal analysis showed that leucine was at the amino terminal position. Protease F hydrolyzes casein and synthetic specific substrates for chymotrypsin and elastase esterases but not for trypsin esterases. It is fully inhibited by phenylmethylsulfonyl fluoride, by chicken ovoinhibitor, and by Chymotrypsin Inhibitor I from potatoes but not by the trypsin-chymotrypsin inhibitors from soybeans and chick peas or by tosyl-L-phenylalanine chloromethyl ketone. The enzyme is stable at room temperature and in the cold, it is not affected by dialysis or by freezing and thawing, but it is inactivated during freeze-drying. The circular dichroism spectra of Protease F indicate an approximate 20% alpha-helix content of the enzyme with a considerable similarity to those of subtilisin, elastase, and beta-trypsin. The relatively low molecular weight of Protease F, the absence of intrachain disulfide bridges, and the fact that it is inhibited by several, but not all, chymotrypsin inhibitors suggest that it may differ phylogenetically from the known serine proteases.

Amino Acids↗
PubMed1982

The effect of dietary raw and autoclaved soya-bean protein fractions on growth, pancreatic enlargement and pancreatic enzymes in rats.

1. Raw soya-bean meal (RS) was fractionated into soya-bean lyophilized extract (SLE), soya-bean lyophilized residue (SLR), acid-precipitated proteins (APP) and whey proteins. 2. Trypsin (EC 3. 4. 21. 4)and chymotrypsin (EC 3. 4. 21.1) inhibitors (TI) were soluble at pH 8 and remained soluble after the extract was acidified to pH 4.4 Except for whey, heating abolished, almost totally, their inhibiting activity. 3. Feeding SLE diet (high TI content) and APP diet (low TI content) resulted in growth depression below the RS level. Feeding the SLR diet resulted in an optimal growth. Feeding diets containing heated fractions improved the growth rate though not to the level observed with heated RS (HS) diet. 4. RS, SLE, APP and whey diets produced similar pancreatic enlargement which could be totally (RS, whey) or partially (SLE, APP) abolished by heating. 5. Feeding the RS diet reduced pancreatic amylase content. The factor responsible for this effect cofractionated with SLE and whey proteins. 6. Two groups of factors in the various diets were probably responsible for the elevation in pancreatic proteases. The first group were the heat-labile factors present in RS, SLE and whey whereas the second group resisted the heat treatment and were found in APP and SLR. 7. The results suggest that for optimal growth rate of rats, heat treatment should be given to the unfractionated soya-bean proteins rather than to the isolated fractions. The results further indicated that TI are not the only factors that can lead to pancreatic enlargement and changes in pancreatic enzymes composition.

Animals↗
PubMed1980

Circular dichroism spectra of cleavage fragments of soybean trypsin-chymotrypsin inhibitor.

Circular dichroism spectra of biologically active fragments of Bowman-Birk soybean inhibitor have been determined in acidic, neutral, and alkaline conditions. Neither fragment showed evidence of alpha-helix or beta-structure. Negative dichroism above 260nm has been assigned in both fragments primarily to disulfide bonds, with a minor contribution from tyrosine in a hydrophilic environment. The individual spectra of these fragments, and their sum between 230 and 340 nm have been compared with the spectra of the intact inhibitor and several structurally related proteins. Possible interactions which may give rise to CD bands in this region are discussed.

Chymotrypsin↗
PubMed1979

Trypsin and chymotrypsin inhibitor from chick peas. Selective chemical modifications of the inhibitor and isolation of two isoinhibitors.

The trypsin and chymotrypsin inhibitor from chick peas (CI) is stable in HCl 0.001 M -- 0.01 M and in KOH 0.01 M -- 0.05 M even after 24 h. Increased KOH concentrations decrease considerably the inhibitory activity already after 1 h. Maleyation and succinylation of the inhibitor resulted in almost full loss of its trypsin-inhibitory activity but had no effect on the chymotrypsin-inhibitory activity. A series of modifications directed towards tyrosyl residues showed that iodination influenced only the chymotrypsin-inhibitory activity; however, nitration and arsanilation affected not only the chymotrypsin-inhibitory activity but also the trypsin-inhibitory activity. Treatment of the inhibitor with CNBr and chloramine T resulted only in a decrease in the chymotrypsin-inhibitory activity indicating that the only methionine is involved in the chymotrypsin-inhibitory activity. When CI-fragment A, previously treated with trypsin at pH 3.75, was further treated with carboxypeptidase B, a release of three lysyl residues per mole protein was found. CI was separated by equilibrium chromatography on SP-Sephadex column into two isoinhibitors, CII and CIII, respectively. Both inhibited trypsin and chymotrypsin with the same specific activity as CI. They differed from each other only in a glutamyl, aspartyl, glycyl and alanyl residue.

Amino Acids↗
PubMed1978

Two fragments from fibrinolysin digests of ovine prolactin: characterization and recombination to generate full immunoreactivity.

The action of fibrinolysin (plasmin; EC 3.4.21.7) on ovine prolactin has been investigated. It was found that the enzyme selectively cleaves the bond between Met-53 and Ala-54. The two fragments, PRL-(1-53) and PRL-(54-199), have been purified and characterized. A recombinant molecule has been obtained by noncovalent interaction of PRL-(1-53) and PRL-(54-199). The recombined protein behaves nearly identically to the parent hormone in circular dichroism spectra and exclusion chromatography. The recombinant possesses full immunoreactivity, as revealed by gel double-diffusion and complement fixation. However, the recombined protein exhibits low prolactin activity in the pigeon crop-sac test.

Amino Acids↗
PubMed1978

Circular dichroism spectra of isolated soybean and chickpea trypsin-chymotrypsin inhibitors.

Circular dichroism spectra of trypsin-chymotrypsin inhibitors from soybeans and chickpeas have been determined in acidic, neutral and highly alkaline solutions. Neither protein contains alpha-helix although a small amount of beta-structure cannot be excluded. Negative dichroism above 250 nm has been assigned largely to disulfide bonds in both molecules with neither showing evidence for tyrosine residues buried in hydrophobic regions. The spectra of these inhibitors between 230 and 250 nm have been compared with the spectra of a number of structurally related proteins suggesting that previous interpretations of this region may have been incomplete or incorrect.

Chymotrypsin↗
PubMed1977

Elastases from human and canine granulocytes, II. Interaction with protease inhibitors of animal, plant, and microbial origin.

Inhibitors of animal, plant, and microbial origin were tested against human and canine granulocytic elastases. The trypsin-chymotrypsin inhibitors from dog submandibular glands, from soybeans (Bowman-Birk) and from chickpeas show strong interaction with these proteases (Ki = 10(-8) - 10(-9)M). The trypsin-kallikrein inactivator of bovine organs (Trasylol) is not active against granulocytic elastases or against human granulocytic cathepsin G. Elastatinal, a specific inhibitor of elastases, isolated from actinomycetes (Streptomyces griseoruber), forms stable complexes with elastase from human (Ki = 6.2 X 10(-6)M) and canine granulocytes (Ki = 1.1 X 10(-6)M). A possible therapeutic application of these inhibitors for the inactivation of granulocytic proteases, which are able to degrade connective tissue in different pathological states, is discussed.

Animals↗
PubMed1976

A trypsin and chymotrypsin inhibitor from chick peas (Cicer arietinum).

1. A trypsin and chymotrypsin inhibitor was isolated by extraction of chick-pea meal at pH8.3, followed by (NH4)2SO4 precipitation and successive column chromatography on CM-cellulose and calcium phosphate (hydroxyapatite). 2. The inhibitor was pure by polyacrylamide-gel and cellulose acetate electrophoresis and by isoelectric focusing in polyacrylamide gels. 3. The inhibitor had a molecular weight of approx. 10000 as determined by ultracentrifugation and by polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate. A molecular weight of 8300 was resolved from its amino acid composition. 4. The inhibitor formed complexes with trypsin and chymotrypsin at molar ratios of 1:1. 5. Limited proteolysis of the inhibitor with trypsin at pH3.75 resulted in hydrolysis of a single-Lys-X-bond and in consequent loss of 85% of the trypsin inhibitory activity and 60% of the chymotrypsin inhibitory activity. Limited proteolysis of the inhibitor with chymotrypsin at pH3.75 resulted in hydrolysis of a single-Tyr-X-bond and in consequent loss of 70% of the trypsin inhibitory activity and in complete loss of the chymotrypsin inhibitory activity. 6. Cleavage of the inhibitor with CNBr followed by pepsin and consequent separation of the products on a Bio Gel P-10 column, yielded two active fragments, A and B. Fragment A inhibited trypsin but not chymotrypsin, and fragment B inhibited chymotrypsin but not trypsin. The specific trypsin inhibitory activity, on a molar ratio, of fragment A was twice that of the native inhibitor, suggesting the unmasking of another trypsin inhibitory site as a result of the cleavage. On the other hand, the specific chymotrypsin inhibitory activity of fragment B was about one-half of that of the native inhibitor, indicating the occurrence of a possible conformational change.

Amino Acids↗