INCORPORATION OF AMINO ACIDS INTO CHICK PROTEINS DURING EMBRYONIC GROWTH

1965 ◽  
Vol 43 (7) ◽  
pp. 1099-1110 ◽  
Author(s):  
Margaret P. Taussig
Keyword(s):  
Amino Acids ◽  
Proteolytic Activity ◽  
Specific Activity ◽  
Sharp Decrease ◽  
Embryonic Growth

Radioactive glycine and methionine were injected into the yolk of 4-day-old embryonated eggs. There was a rapid increase in total incorporated radioactivity with both amino acids. The specific activity of the protein increased until the seventh day, after which time there was a sharp decrease. Radioactivity from DL-methionine-S35 was incorporated in larger amounts than from glycine-2-C14. An increase in the proteolytic activity of the yolk with age was demonstrated.

A New Method for Plasminogen Standardization

1968 ◽  
Vol 20 (03/04) ◽  
pp. 548-554
Author(s):  
J Gajewski ◽  
G Markus
Keyword(s):  
Proteolytic Activity ◽  
Specific Activity ◽  
Molar Ratio ◽  
Sharp Transition ◽  
Equivalence Point ◽  
Constant Amount ◽  
Residual Level ◽  
Activity Measurements ◽  
Complete Saturation ◽  
Human Plasminogen

SummaryA method for the standardization of human plasminogen is proposed, based on the stoichiometric interaction between plasminogen and streptokinase, resulting in inhibition of proteolytic activity. Activation of a constant amount of plasminogen with increasing amounts of streptokinase yields linearly decreasing activities, as a function of streptokinase, with a sharp transition to a constant residual level. The point of transition corresponds to complete saturation of plasmin with streptokinase in a 1:1 molar ratio, and is therefore a measure of the amount of plasminogen present initially, in terms of streptokinase equivalents. The equivalence point is independent of the kind of protein substrate used, buffer, pH, length of digestion and, within limits, temperature. The method, therefore, is not subject to the variations commonly encountered in the usual determination based on specific activity measurements.

scholarly journals A multi-enzyme cascade for efficient production of d-p-hydroxyphenylglycine from l-tyrosine

2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Xu Tan ◽  
Sheng Zhang ◽  
Wei Song ◽  
Jia Liu ◽  
Cong Gao ◽  
...  
Keyword(s):  
Amino Acids ◽  
Specific Activity ◽  
Enantiomeric Excess ◽  
Efficient Production ◽  
Enzyme Cascade ◽  
Rate Limiting Step ◽  
Limiting Step ◽  
Rotation Strategy ◽  
Rate Limiting

AbstractIn this study, a four-enzyme cascade pathway was developed and reconstructed in vivo for the production of d-p-hydroxyphenylglycine (D-HPG), a valuable intermediate used to produce β-lactam antibiotics and in fine-chemical synthesis, from l-tyrosine. In this pathway, catalytic conversion of the intermediate 4-hydroxyphenylglyoxalate by meso-diaminopimelate dehydrogenase from Corynebacterium glutamicum (CgDAPDH) was identified as the rate-limiting step, followed by application of a mechanism-guided “conformation rotation” strategy to decrease the hydride-transfer distance d(C6HDAP−C4NNADP) and increase CgDAPDH activity. Introduction of the best variant generated by protein engineering (CgDAPDHBC621/D120S/W144S/I169P with 5.32 ± 0.85 U·mg−1 specific activity) into the designed pathway resulted in a D-HPG titer of 42.69 g/L from 50-g/L l-tyrosine in 24 h, with 92.5% conversion, 71.5% isolated yield, and > 99% enantiomeric excess in a 3-L fermenter. This four-enzyme cascade provides an efficient enzymatic approach for the industrial production of D-HPG from cheap amino acids.

scholarly journals Utilization in vivo of glucose and volatile fatty acids by sheep brain for the synthesis of acidic amino acids

1966 ◽  
Vol 101 (3) ◽  
pp. 591-597 ◽  
Author(s):  
R M O'Neal ◽  
R E Koeppe ◽  
E I Williams
Keyword(s):  
Amino Acids ◽  
Fatty Acids ◽  
Glutamic Acid ◽  
Aspartic Acid ◽  
Free Amino Acids ◽  
Free Amino ◽  
Specific Activity ◽  
Tricarboxylic Acid Cycle ◽  
Sheep Brain ◽  
The Brain

1. Free glutamic acid, aspartic acid, glutamic acid from glutamine and, in some instances, the glutamic acid from glutathione and the aspartic acid from N-acetyl-aspartic acid were isolated from the brains of sheep and assayed for radioactivity after intravenous injection of [2-(14)C]glucose, [1-(14)C]acetate, [1-(14)C]butyrate or [2-(14)C]propionate. These brain components were also isolated and analysed from rats that had been given [2-(14)C]propionate. The results indicate that, as in rat brain, glucose is by far the best precursor of the free amino acids of sheep brain. 2. Degradation of the glutamate of brain yielded labelling patterns consistent with the proposal that the major route of pyruvate metabolism in brain is via acetyl-CoA, and that the short-chain fatty acids enter the brain without prior metabolism by other tissue and are metabolized in brain via the tricarboxylic acid cycle. 3. When labelled glucose was used as a precursor, glutamate always had a higher specific activity than glutamine; when labelled fatty acids were used, the reverse was true. These findings add support and complexity to the concept of the metabolic; compartmentation' of the free amino acids of brain. 4. The results from experiments with labelled propionate strongly suggest that brain metabolizes propionate via succinate and that this metabolic route may be a limited but important source of dicarboxylic acids in the brain.

scholarly journals ω-Amino Acid:Pyruvate Transaminase from Alcaligenes denitrificans Y2k-2: a New Catalyst for Kinetic Resolution of β-Amino Acids and Amines

2004 ◽  
Vol 70 (4) ◽  
pp. 2529-2534 ◽  
Author(s):  
Hyungdon Yun ◽  
Seongyop Lim ◽  
Byung-Kwan Cho ◽  
Byung-Gee Kim
Keyword(s):  
Amino Acids ◽  
Kinetic Resolution ◽  
Specific Activity ◽  
Expression System ◽  
Enantiomeric Excess ◽  
Selective Enrichment ◽  
Alcaligenes Denitrificans ◽  
Keto Acids ◽  
Optically Pure ◽  
High Stereoselectivity

ABSTRACT Alcaligenes denitrificans Y2k-2 was obtained by selective enrichment followed by screening from soil samples, which showed ω-amino acid:pyruvate transaminase activity, to kinetically resolve aliphatic β-amino acid, and the corresponding structural gene (aptA) was cloned. The gene was functionally expressed in Escherichia coli BL21 by using an isopropyl-β-d-thiogalactopyranoside (IPTG)-inducible pET expression system (9.6 U/mg), and the recombinant AptA was purified to show a specific activity of 77.2 U/mg for l-β-amino-n-butyric acid (l-β-ABA). The enzyme converts various β-amino acids and amines to the corresponding β-keto acids and ketones by using pyruvate as an amine acceptor. The apparent Km and V max for l-β-ABA were 56 mM and 500 U/mg, respectively, in the presence of 10 mM pyruvate. In the presence of 10 mM l-β-ABA, the apparent Km and V max for pyruvate were 11 mM and 370 U/mg, respectively. The enzyme exhibits high stereoselectivity (E > 80) in the kinetic resolution of 50 mM d,l-β-ABA, producing optically pure d-β-ABA (99% enantiomeric excess) with 53% conversion.

Effects of nonsulfur and sulfur amino acids on the regulation of hepatic enzymes of cysteine metabolism

1999 ◽  
Vol 277 (1) ◽  
pp. E144-E153 ◽  
Author(s):  
Deborah L. Bella ◽  
Christine Hahn ◽  
Martha H. Stipanuk
Keyword(s):  
Amino Acids ◽  
Specific Activity ◽  
Mrna Level ◽  
Basal Diet ◽  
Cysteine Dioxygenase ◽  
Sulfur Amino Acids ◽  
Subunit Mrna ◽  
Cysteine Metabolism ◽  
Glutamylcysteine Synthetase

To determine the role of nonsulfur vs. sulfur amino acids in regulation of cysteine metabolism, rats were fed a basal diet or diets supplemented with a mixture of nonsulfur amino acids (AA), sulfur amino acids (SAA), or both for 3 wk. Hepatic cysteine-sulfinate decarboxylase (CSDC), cysteine dioxygenase (CDO), and γ-glutamylcysteine synthetase (GCS) activity, concentration, and mRNA abundance were measured. Supplementation with AA alone had no effect on any of these measures. Supplementation of the basal diet with SAA, with or without AA, resulted in a higher CDO concentration (32–45 times basal), a lower CSDC mRNA level (49–64% of basal), and a lower GCS-heavy subunit mRNA level (70–76%). The presence of excess SAA and AA together resulted in an additional type of regulation: a lower specific activity of all three enzymes was observed in rats fed diets with an excess of AA and SAA. Both SAA and AA played a role in regulation of these three enzymes of cysteine metabolism, but SAA had the dominant effects, and effects of AA were not observed in the absence of SAA.

scholarly journals Site-directed mutagenesis at aspartate and glutamate residues of xylanase from Bacillus pumilus

1992 ◽  
Vol 288 (1) ◽  
pp. 117-121 ◽  
Author(s):  
E P Ko ◽  
H Akatsuka ◽  
H Moriyama ◽  
A Shinmyo ◽  
Y Hata ◽  
...  
Keyword(s):  
Amino Acids ◽  
Reaction Mechanism ◽  
Amino Acid ◽  
Bacillus Pumilus ◽  
Specific Activity ◽  
Sequence Similarity ◽  
Site Directed Mutagenesis ◽  
Dimensional Structure ◽  
Amino Acid Sequence Similarity ◽  
Directed Mutagenesis

To elucidate the reaction mechanism of xylanase, the identification of amino acids essential for its catalysis is of importance. Studies have indicated the possibility that the reaction mechanism of xylanase is similar to that of hen's egg lysozyme, which involves acidic amino acid residues. On the basis of this assumption, together with the three-dimensional structure of Bacillus pumilus xylanase and its amino acid sequence similarity to other xylanases of different origins, three acidic amino acids, namely Asp-21, Glu-93 and Glu-182, were selected for site-directed mutagenesis. The Asp residue was altered to either Ser or Glu, and the Glu residues to Ser or Asp. The purified mutant xylanases D21E, D21S, E93D, E93S, E182D and E182S showed single protein bands of about 26 kDa on SDS/PAGE. C.d. spectra of these mutant enzymes show no effect on the secondary structure of xylanase, except that of D21E, which shows a little variation. Furthermore, mutations of Glu-93 and Glu-182 resulted in a drastic decrease in the specific activity of xylanase as compared with mutation of Asp-21. On the basis of these results we propose that Glu-93 and Glu-182 are the best candidates for the essential catalytic residues of xylanase.

THE PROTEOLYTIC ACTIVITY OF INSULIN

1967 ◽  
Vol 45 (9) ◽  
pp. 1329-1333
Author(s):  
Michel Page ◽  
Claude Godin
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Proteolytic Activity ◽  
Protein Amino Acids

The action of insulin on hemoglobin at pH 7.5 was studied. Four different methods were used to determine the degree of proteolysis. After mixtures of hemoglobin and insulin were incubated for 2 hours, very little amino acid or peptide material was liberated from the proteins. As many amino acids are liberated from hemoglobin when it is incubated alone under the same conditions. It is concluded that autolysis is responsible for the observed increases in non-protein amino acids and that insulin has no proteolytic activity under the conditions used in this study.

scholarly journals SELECTED PROTEOLYTIC ACTIVITY BY EXTRACTS OF DERMESTID LARVAE

2011 ◽  
Vol 19 (19) ◽  
pp. 79-83
Author(s):  
Lavinel G. IONESCU
Keyword(s):  
Enzyme Activity ◽  
Ammonium Sulfate ◽  
Proteolytic Activity ◽  
Specific Activity ◽  
Proteolytic Enzymes ◽  
High Specific Activity ◽  
Water Extract ◽  
Dermestes Maculatus ◽  
Crude Preparation

The larvae of the Beetle Dermestes maculatus De Geer can subsist on a diet consisting largely of protein. Studies have been undertaken to investigate the nature of proteolytic enzymes. A water extract of the larvae yielded a crude preparation that hydrolyzes gelatin, bide powder, hemoglobin substrate, benzoyl-DL-arginine p-nitroamilide, and glutaryl-L-phenylalanine p-nitroanilide. Enzyme activity was found in a non-dialyzable, heat- and acid0labile portion of the extract yielded two fractions with high specific activity towards gelatin. These are precipitated between 40% to 60% saturation of ammonium sulfate and 60% to 80% saturation. The higher specific activity was observed in the 40%-60% fraction. These results suggest that the larvae of these dermestids contain proteolytic enzymes with actions similar to mammalian trypsin and chymotrypsin. The results also suggest that other proteolytic enzymes may be present as well.

STUDIES ON WHEAT PLANTS USING CARBON-14 LABELLED COMPOUNDS: X. THE INCORPORATION OF GLUTAMIC ACID-l-C14

1959 ◽  
Vol 37 (1) ◽  
pp. 933-936 ◽  
Author(s):  
W. B. McConnell
Keyword(s):  
Amino Acids ◽  
Glutamic Acid ◽  
Specific Activity ◽  
Wheat Plant ◽  
Protein Fractions ◽  
Appreciable Amount ◽  
Kernel Development ◽  
Carbon 14

Glutamic acid-1-C14 was injected into the top internode of wheat stems at a stage of growth when kernel development was rapid (71 days after seeding). The plants were harvested 31 days later when they had matured and the incorporation of carbon-14 studied. About one-third of the carbon-14 administered was found in the upper portions of the mature plants, much of the remaining radioactivity having apparently been respired. About 85% of the carbon-14 recovered was found in the kernel. The protein fractions of these were most radioactive, but an appreciable amount of carbon-14 also appeared in the starch. Glutamic acid had the highest specific activity of the amino acids isolated from the gluten, but proline and arginine were also strongly labelled. Since these three amino acids were labelled predominantly in carbon-1 their close metabolic relationship in the wheat plant seems probable.

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