Acid Carboxypeptidase Research Articles

Purification and Properties of Acid Carboxypeptidase IV fromAspergillus oryzae

Purification and Properties of Acid Carboxypeptidase IV from<i>Aspergillus oryzae</i>

Specificity and mode of action of acid carboxypeptidase from Aspergillus saitoi

1. 1. Acid carboxypeptidase isolated from culture filtrate of Aspergillus saitoi has been investigated for its use in carboxyterminal sequence determination of native insulin, angiotensin I and glucagon at pH 2.5 or 2.2. The examination indicated that after 5 h incubation asparagine, alanine and lysine were liberated from native insulin. The acid carboxypeptidase catalyzed the release of leucine and histidine from the decapeptide angiotensin I. When (Pro-ProGly) 5 and polylysine were incubated separately with the acid carboxypeptidase, no hydrolysis was found. No release of free amino acids by autodigestion of the enzyme was detected after a prolonged period of incubation. 2. 2. The pH dependene of log ( V/ K m ) and log V for the acid carboxypeptidase-catalyzed hydrolysis of a small synthetic peptide, Z-Tyr-Leu, has been determined. Bell-shaped pH-rate profiles were obtained. The main conclusions derived from analysis of the data were that two catalytically-active groups on the enzyme with p K e1 ≅ 2.3 and p K e2 ≅ 4.9 were important in the enzymatic action at the acid end of the pH range. Competitive inhibition by small substrates was found with hydrocinnamic acid, indole-3-propionic acid and 4-phenylbutyric acid. The K i value for hydrocinnamic acid inhibition was 4·10 −4 M.

Production of a New Type of Acid Carboxypeptidase of Molds of the Aspergillus niger Group

The ability of 88 fungi, which had been obtained as high-potency strains for acid proteinase production, to produce a new type of acid carboxypeptidase (having on optimal pH of about 3 for hydrolysis of benzyloxycarbonyl-glutamyltyrosine) in surface koji culture was determined. Among the aspergilli, substantial amounts of this new acid carboxypeptidase were produced by Aspergillus saitoi, A. usamii, A. awamori, A. inuii, and A. niger. Maximum yields of acid carboxypeptidase per gram of substrate were obtained by submerged culture in a medium containing 0.9% defatted soybean and 0.6% wheat bran. However, the maximum enzyme concentration per milliliter was obtained with a medium containing 3% defatted soybean and 2% wheat bran. The terminal pH could be controlled by varying the concentrations of soybean oil meal and wheat bran. The maximum enzyme production was reached after 4 days or more at 30 C.

The Effect of Galactosamine on the Activity of Lysosomal Peptidehydrolases of the Liver Depending on the Age of Rats

In order to elucidate the role of lysosomes during the development of hepatitis induced by D-galactosamine, the activities of cathepsin A, cathepsin D, and acid carboxypeptidase are measured in the liver after administration of a single dose of 375 mg D-galactosamine HCI per kg body weight This dose is able to induce hepatitis in young and adult, but not in newborn rats up to 4 weeks of age. Concommitantly, changes in lysosomal enzyme activities are not observed in the livers of newborn rats but in those of young and adult rats.

Esterolytic Activity of Acid Carboxypeptidase from Aspergillus saitoi

Journal Article Esterolytic Activity of Acid Carboxypeptidase from Aspergillus saitoi Get access Eiji Ichishima, Eiji Ichishima Laboratory of Microbiology and Enzymology, Tokyo Nōkō University, Fuchu, Tokyo 183 Search for other works by this author on: Oxford Academic Google Scholar Kenji Yomogida Kenji Yomogida Laboratory of Microbiology and Enzymology, Tokyo Nōkō University, Fuchu, Tokyo 183 Tokyo Metropolitan Inspection Laboratories for Feed and Fertilizers, Marunouchi, Tokyo 101. Search for other works by this author on: Oxford Academic Google Scholar Agricultural and Biological Chemistry, Volume 37, Issue 3, 1 March 1973, Pages 693–694, https://doi.org/10.1080/00021369.1973.10860734 Published: 01 March 1973 Article history Received: 11 November 1972 Published: 01 March 1973

Purification and Properties of Acid Carboxypeptidase IV from Aspergillus oryzae

Acid carboxypeptidase IV from Aspergillus oryzae was purified from the rivanol precipitable fraction by column chromatography on DEAE-cellulose, DEAE-Sephadex A–50, hydroxylapatite and P-cellulose and gel filtration through Sephadex G–100. The optimum pH is at pH 3.0 for carbobenzoxy-l-glutamyl-l-tyrosine. The enzyme activity was inhibited by sulfhydryl reagents and diisopropylphosphorofluoridate, but was not inhibited by metal chelating agents. The molecular weight of the enzyme was estimated to be about 43,000 by gel filtration method.

Comparative study on enzymatic properties of acid carboxypeptidase of molds of the genus Aspergillus.

Comparative study on enzymatic properties of acid carboxypeptidase of molds of the genus Aspergillus.

Purification and Properties of Acid Carboxypeptidase II fromAspergillus oryzae

Acid carboxypeptidase II from Aspergillus oryzae was purified from the rivanol non-precipitated fraction. The purified enzyme was homogeneous on polyacrylamide gel disc electrophoresis. The optimum activity of the enzyme lay at pH 3.0 for carbobenzoxy-L-glutamyl-l-tyrosine. The enzyme was inhibited by diisopropylphosphorofluoridate and SH reagents such as p-chloromercuribenzoate and monoiodoacetate, but not by such metal chelating agents as ethylenediaminetetraacetate, α, α′-dipyridyl and o-phenanthroline. The molecular weight of the enzyme was estimated to be about 105,000.

Purification and Properties of Acid Carboxypeptidase III fromAspergillus oryzae

Purification and Properties of Acid Carboxypeptidase III from<i>Aspergillus oryzae</i>

Purification and Properties of Acid Carboxypeptidase I fromAspergillus oryzae

Acid carboxypeptidase III from Aspergillus oryzae was purified from the rivanol non-precipitated fraction. The optimum activity of the enzyme occurred at pH 3.0 for carbobenzoxy-l-glutamyl-l-tyrosine. The enzyme was inhibited by diisopropylphosphorofluoridate and SH reagents such as p-chloromercuribenzoate and monoiodoacetate, but not by such metal chelating agents as ethylenediaminetetraacetate, αα′-dipyridyl and o-phenanthroline. The molecular weight of the enzyme was estimated to be about 61,000. The enzyme hydrolyzed the peptides that possess masked or bulky N-terminal.

Anticoagulant Peptide Obtained from Fibrinogen Degradation Products by Plasmin. II

The amino acid sequence of a decapeptide which had an inhibitory effect on the blood coagulation was determined by Edman degradation, dansyl-Edman procedure, and hydrolysis by acid carboxypeptidase. The amino acid sequence of this peptide was determined as follows:Ser-Gln-Leu-Gln-Glu-Ala-Pro-Leu-Glu-Lys.

Zur Auftrennung und Charakterisierung proteolytischer Enzyme aus Euglena gracilis

Summary To test the activities of proteolytic enzymes in Euglena gracilis Klebs (1224 — 5/25, strain Z) some procedures of cell destruction were compared. Destruction with Alcoa A 305 had been most suitable. In separation of the proteolytic enzyme spectrum Sephadex G-200 showed the best results. The following activities were differentiated: 1. Amino acid arylamidase I (splits Leu-NA better than Ala-NA, except on the pH-optimum (for Leu 7,3, for Ala 7,7) and better than Arg-NA (pH-optimum 7,1). 2. L-Leucine aminopeptidase (splits Leu-NH2 and Leu-Gly-Gly; activated by Mn2+). 3. Amino acid arylamidase II (less activity than I, splits Ala-NA always better than Leu-NA better than Arg-NA). 4. L-aminoacid acylase (splits acetyl- and chloracetyl-aminoacids on pH 8,2—8,4; the enzyme is independent of metal ions; molecular weight about 78.000). 5. L-amino tri(poly)peptidase (splits on pH 7.8 Leu-Gly-Gly and similar tripeptides as well as Gly-Gly-D-Leu; inhibition by Mn2+). 6. Acid carboxypeptidase (splits Z-L-dipeptides or tripeptides on pH 4,5 —5,7). 7. L-aminoacyl-L-aminoacid-hydrolase (dipeptidase).

Purification and Properties of Acid Carboxypeptidase III from Aspergillus oryzae

Purification and Properties of Acid Carboxypeptidase III from Aspergillus oryzae

Purification and characterization of a new type of acid carboxypeptidase from Aspergillus

1. 1.|This paper reports the purification and characterization of a new type of acid carboxypeptidase from Aspergillus. The highly purified enzyme was found to be essentially homogeneous by such criteria as sedimentation in the ultracentrifuge and disc electrophoreses on polyacrylamide gel at pH 9.5, 8.0 and 2.3. 2. 2.|The enzyme has a pH optimum at pH 3.1 for Z-Glu-Tyr, pH 3.5 for Z-Tyr-Leu, pH 3.2 for Z-Gly-Pro-Leu-Gly, and pH 3.5 for Bz-Gly-Lys. The K m and ν max values for Z-Glu-Tyr at pH 3.1 and 30° are 1.25 · 10 −3 M and 1.9 · 10 4 μmoles tyrosine per min for A 280 nm, the values for Z-Tyr-Leu at pH 3.5 and 30° are 1.0–10 −3 M and 1.64 · 10 5 μmoles leucine/min for A 280 nm, and the values for Bz-Gly-Lys at pH 3.5 and 30° are 4.0 · 10 −3 M and 5.5 · 10 2 μmoles lysine per min for A 280 nm. The specificity of the enzyme was investigated for a wider range of substrates at the acid end of the pH spectrum and the carboxypeptidase-like activity was confirmed. 3. 3.|According to the Yphantis' procedure, the molecular weight of the enzyme at 0.2 ionic strength was found to be 139 000. The sedimentation constant was determined to be 7.3 S. According to the gel filtration method, molecular weight values 155 000 for the larger form or polymer and 51 000 for the smaller form or monomer were obtained in the absence of NaCl, and a molecular weight value of 135 000 was obtained in the presence of 0.2 M NaCl. 4. 4.|The enzymatic activity was inhibited by DFP at pH 6.0 and 0°, and was not affected by EDTA at pH 5.2 and 5°.

Purification and Properties of Acid Carboxypeptidase I from Aspergillus oryzae

Purification and Properties of Acid Carboxypeptidase I from Aspergillus oryzae

Purification and Properties of Acid Carboxypeptidase II from Aspergillus oryzae

Acid carboxypeptidase II from Aspergillas oryzae was purified from the rivanol nonprecipitated fraction. The purified enzyme was homogeneous on polyacrylamide gel disc electrophoresis. The optimum activity of the enzyme lay at pH 3.0 for carbobenzoxy-L-glutamyl-L-tyrosine. The enzyme was inhibited by diisopropylphosphorofluoridate and SH reagents such as p-chloromercuribenzoate and monoiodoacetate, but not by such metal chelating agents as ethylenediaminetetraacetate, α, α'-dipyridyl and o-phenanthroline. The molecular weight of the enzyme was estimated to be about 105, 000.

APPLYING PROTEOLYTIC ENZYMES ON SOYBEAN. 6. Deodorization Effect of Aspergillopeptidase A and Debittering Effect of Aspergillus Acid Carboxypeptidase

SUMMARY— A proteolyzate obtained by treating an isolated soybean protein preparation with Molsin, a crude preparation of aspergillopeptidase A (APase A), was less bitter and contained larger amounts of free amino acids, especially hydrophobic amino acids. A proteolyzate obtained by treating this protein preparation with crystallized APase A was much more bitter and contained smailer amounts of free amino acids, mainly consisting of hydrophilic amino acids. The latter was richer in peptides than the former, bearing hydrophobic amino acid residues near the C‐termini. Difference in N‐terminal amino acid composition apparently has not been found between the 2 proteolyzates. These results indicate that Molsin per se contains a certain carboxypeptidase which decomposes the C‐terminal structures and, consequently, lessens the bitterness (debittering effect). This carboxypeptidase was found to be identical with aspergillus acid carboxypeptidase (AACPase). Abase A, as well as MO/sin, was effective in removing odor ants, i.e., n‐hexanal, n‐hexanol and n‐heptanol, from the isolated soybean protein preparation Ideodorization effect). AACPase seemed to have no deodorization effect. A method was suggested to prepare a deodorized and debittered proteolyzate by a combination use of APase A and AACPase.

Applying Proteolytic Enzymes on Soybean

The cold-insoluble fraction of soy protein was hydrolyzed with pepsin under usual conditions and the hydrolyzate was dialyzed. The nondialyzable part was treated with aqueous ethanol to obtain a 50% ethanol-soluble and 90% ethanol-insoluble fraction. From this fraction, a bitter tetracosapeptide was separated with the aid of column chromatography using Sephadex G-10 and DEAE Sephadex A-25, thin layer chromatography using silica gel G and cellulose powder, and paper electrophoresis. The partial structure of the bitter tetracosapeptide was proposed as follows: H•Phe-(Arg, Asp2, Glu2, Gly, Ile, Leu, Lys2, Pro, Ser, Thr)-Trp-(Ala, Arg, Asp, Gly, Val)-Gln-Tyr-Phe-Leu•OH. Pepsin hydrolyzed this peptide to lessen its bitterness, whereas trypsin was ineffective in lessening the bitterness. Either carboxypetidase A or aspergillus acid carboxypeptidase was effective in debittering. N-Bromosuccinimide treatment on this peptide produced a nonapeptide bearing the intact C-terminal structure, which was found to be still bitter. However, further degradation of the bitter nonapeptide with the carboxypeptidase A or the aspergillus acid carboxypeptidase from its C-terminal side lenssened the bitterness. These results indicate that the bitterness of the bitter tetracosapeptide relates intimately to the hydrophobic C-terminal sequence, -Tyr-Phe-Leu•OH. Experimental results concerning chemical modification on this tetracosapeptide and also concerning some synthesized model oligopeptides supported the above conclusion.

Acid Carboxypeptidases: Their Occurrence in Plants, Intracellular Distribution and Possible Function

Saure Carboxypeptidase, welche Phenylalanin von CbO-Leu-Phe-OH bei pH 5,3 abspaltet, ist ein in höheren Angiospermen weit verbreitetes Enzym. Es scheint dagegen in primitiven Angiospermen (Fagus und Magnolia) sowie in Gymnospermen und Kryptogamen (Ausnahme: Equisetum) zu fehlen. Bei Zitrone ist die Aktivität saurer Carboxypeptidase von Natur und Alter der extrahierten Gewebe abhängig (Exocarp und Endocarp der Früchte, Blätter). In Zellen der Wurzelspitzen von Maiskeimlingen ist das Enzym in meristematischen Vakuolen lokalisiert. Funktionell ist die saure Carboxypeptidase als lysosomales Enzym, welches am intrazellulären Proteinabbau teilnimmt, anzusprechen.

Proteolytic activity of human gastric mucosa. I.

1. In human gastric mucosa besides peptic action, are found cathepsin action (pH 4.0-5.0, casein, egg albumin and serum albumin ) and catheptic _??_ pepton splitting action (cathopeptonase). And casein is also at pH 8.5 hydrolized (72 hrs.). 2. I have succeeded also with human gastric mucosa to separate the pepsin action from the cathepsin action, and also the cathopeptonase action from the erepsin action by means of acid and alkaline treatment respectively according to the methods of Utzino and Sakai.3) 3. In human gastric mucosa halogen-acylase action (pH 7.8, chlor-acetyl-1-leucin and chloracetyl-1-phenylalanin) is found, but carboxy-peptidase action (pH 7.8, benzoyl-diglycin, with kinase) or hippuricase action (pH 7.8, benzoyl-glycin) not. The hydrolysis of chlor-acetyl- _??_ diglycin at pH 7.8 by gastric mucosa from ulcer stomach is certain extent recognized. 4. Neither acid carboxy-peptidase action nor. acid acyldipeptidase action (pH 4.2-4.6, benzoyl-diglycin, chloracetyl-diglycin) despite of the cystein activation is still yet found. The hydrolysis of diglycin, chlor-acetyl-l-leucin, chloracetyl-phenylalnin and benzoyl-glycin at pH 4.5 with cystein, all of these are negative.