REACTION OF FORMALDEHYDE AND THIOFORMALDEHYDE WITH MERCURIPAPAIN

1965 ◽  
Vol 43 (7) ◽  
pp. 1171-1177 ◽  
Author(s):  
W. A. Darlington ◽  
L. Keay
Keyword(s):  
Enzyme Activity ◽  
Amino Acid ◽  
Functional Groups ◽  
Tertiary Structure ◽  
Colorimetric Assay ◽  
Amino Groups ◽  
Full Activity ◽  
Amino Acid Side Chains ◽  
Lysine Residues ◽  
Secondary And Tertiary Structure

In a colorimetric assay with benzoyl-DL-arginine p-nitroanilide, acetylated and benzoylated papains retain full activity. Thus the ε-amino groups of the lysine residues are not required for enzyme activity. Intramolecular crosslinking of an enzyme could in theory stabilize secondary and tertiary structure and oppose denaturation. Thioformaldehyde is much more reactive with mercuripapain than formaldehyde, incorporating much more readily into the enzyme at equivalent concentrations. Incorporation is extensive, however, on reactive functional groups on the amino acid side chains, since acetylation decreased the incorporation markedly. In no case was there evidence of heat stabilization.

A Strategy for Thioamide Incorporation During Fmoc Solid-Phase Peptide Synthesis with Robust Stereochemical Integrity

2019 ◽  
Author(s):  
luis camacho III ◽  
Bryan J. Lampkin ◽  
Brett VanVeller
Keyword(s):  
Amino Acid ◽  
Functional Groups ◽  
Peptide Synthesis ◽  
Solid Phase ◽  
Solid Phase Peptide Synthesis ◽  
Side Chains ◽  
Amino Acid Side Chains ◽  
Peptide Elongation

We describe a method to protect the sensitive stereochemistry of the thioamide—in analogy to the protection of the functional groups of amino acid side chains—in order to preserve the thioamide moiety during peptide elongation.<br>

Ringöffnungsreaktionen an bioreaktiven Lactamsystemen / Ring Opening Reactions of Bioreactive Lactam Systems

1987 ◽  
Vol 42 (5) ◽  
pp. 603-612 ◽  
Author(s):  
Hermann Frister ◽  
Eckhard Schlimme
Keyword(s):  
Amino Acid ◽  
Ring Opening ◽  
Side Chains ◽  
Amino Groups ◽  
Ring Opening Reaction ◽  
Ring Opening Reactions ◽  
Amino Acid Side Chains ◽  
Tin Tetrachloride ◽  
Model Reactions

Abstract 1-β-ᴅ-Ribofuranosylpyrrolidin-2,5-dione (9) was synthesized by ribosylation of N-silylated succinimide (7) with 1,2,3,5-tetra-O-acetyl-β-ᴅ-ribofuranose in acetonitril in the presence of tin tetrachloride. The compounds 9, 1-β-ᴅ-ribofuranosyl-l-H-pyrrol-2,5-dione (5) and N-methyl- maleinimide (2) were converted with ammonia to the ring-opened components 16. 14 and 15. The bioreactivity of the N-maleinimide derivatives 2 and 5 with respect to addition and ring-opening reactions with amino acid side chains containing either thiol or amino groups was shown in model reactions with glutathion (compds. 17,18) and lysine (compds. 19, 20). The ring opening reaction of 3-methyl-3-phenyl-1-β-ᴅ-ribofuranosylpyrrolidin-2,5-dione (11) with lysine yields 21, thus demonstrating the possibility of glycosuccinylation of amino groups in proteins.

scholarly journals Rapid removal of acetimidoyl groups from proteins and peptides. Applications to primary structure determination

1981 ◽  
Vol 199 (2) ◽  
pp. 335-340 ◽  
Author(s):  
G C Dubois ◽  
E A Robinson ◽  
J K Inman ◽  
R N Perham ◽  
E Appella
Keyword(s):  
Amino Acid ◽  
Primary Structure ◽  
Half Life ◽  
Structural Studies ◽  
Amino Groups ◽  
Mouse Immunoglobulin ◽  
Lysine Residues ◽  
Rapid Removal ◽  
Proteins And Peptides

Methylamine buffers can be used for the rapid quantitative removal of acetimidoyl groups from proteins and peptides modified by treatment with ethyl or methyl acetimidate. The half-life for displacement of acetimidoyl groups from fully amidinated proteins incubated in 3.44 M-methylamine/HCl buffer at pH 11.5 and 25 degrees C was approx. 26 min; this half life is 29 times less than that observed in ammonia/HCl buffer under the same conditions of pH and amine concentration. Incubation of acetimidated proteins with methylamine for 4 h resulted in greater than 95% removal of acetimidoyl groups. No deleterious effects on primary structure were detected by amino acid analysis or by automated Edman degradation. Reversible amidination of lysine residues, in conjunction with tryptic digestion, has been successfully applied to the determination of the amino acid sequence of an acetimidated mouse immunoglobulin heavy chain peptide. The regeneration of amino groups in amidinated proteins and peptides by methylaminolysis makes amidination a valuable alternative to citraconoylation and maleoylation in structural studies.

scholarly journals Ionic properties of an essential histidine residue in pig heart lactate dehydrogenase

1973 ◽  
Vol 131 (4) ◽  
pp. 729-738 ◽  
Author(s):  
J. John Holbrook ◽  
V. Ann Ingram
Keyword(s):  
Enzyme Activity ◽  
Amino Acid ◽  
Lactate Dehydrogenase ◽  
Side Chains ◽  
Histidine Residue ◽  
Acylating Agent ◽  
Diethyl Pyrocarbonate ◽  
Substrate Analogues ◽  
Amino Acid Side Chains

1. Pig heart lactate dehydrogenase is inhibited by addition of one equivalent of diethyl pyrocarbonate. The inhibition is due to the acylation of a unique histidine residue which is 10-fold more reactive than free histidine. No other amino acid side chains are modified. 2. The carbethoxyhistidine residue slowly decomposes and the enzyme activity reappears. 3. The essential histidine residue is only slightly protected by the presence of NADH but is completely protected when substrate and substrate analogues bind to the enzyme–NADH complex. The protection is interpreted in terms of a model in which substrates can only bind to the enzyme in which the histidine residue is protonated and is thus not available for reaction with the acylating agent. 4. The apparent pKa of the histidine residue in the apoenzyme is 6.8±0.2. In the enzyme–NADH complex it is 6.7±0.2. 5. Acylated enzyme binds NADH with unchanged affinity. The enzyme is inhibited because substrates and substrate analogues cannot bind at the acylated histidine residue in the enzyme–NADH complex.

THE CHEMICAL MODIFICATION OF CHYMOTRYPSIN

1964 ◽  
Vol 42 (6) ◽  
pp. 695-714 ◽  
Author(s):  
G. H. Dixon ◽  
H. Schachter
Keyword(s):  
Enzyme Activity ◽  
Amino Acid ◽  
Chemical Modification ◽  
Active Site ◽  
Specific Binding ◽  
Bond Breaking ◽  
Kinetic Properties ◽  
Side Chains ◽  
Amino Acid Side Chains

Chemical modification of chymotrypsin has led to the identification of several amino acid side-chains which are probably constituents of the active site of the enzyme. A single seryl and a single histidyl residue appear to cooperate in catalyzing the bond-breaking process while one or more tryptophanyl residues may be involved in the specific binding of substrate. Neither of the two methionyl residues is essential for enzyme activity although changes in kinetic properties occur when they are modified by oxidation or alkylation.

Detecting ACCase-targeting herbicides effect on ACCase activity utilizing a malachite green colorimetric functional assay

Weed Science ◽  
2021 ◽  
pp. 1-18
Author(s):  
Suma Basak ◽  
Douglas Goodwin ◽  
Jahangir Alam ◽  
James Harris ◽  
Jinesh D. Patel ◽  
...  
Keyword(s):  
Enzyme Activity ◽  
Amino Acid ◽  
Malachite Green ◽  
Amino Acid Substitution ◽  
Dose Response ◽  
Colorimetric Assay ◽  
Functional Assay ◽  
Acetyl Coenzyme A ◽  
Order Of Magnitude ◽  
Acetyl Coenzyme A Carboxylase

Research was conducted to evaluate acetyl-Coenzyme A carboxylase (ACCase) enzyme activity using a functional malachite green colorimetric assay previously identified as resistant to sethoxydim, and select aryloxyphenoxypropionate (FOPs) herbicides, fenoxaprop, and fluazifop. Two resistant southern crabgrass [Digitaria ciliaris (Retz.) Koeler] biotypes, R1 and R2, containing an Ile-1781-Leu amino acid substitution and previously identified as resistant to sethoxydim, pinoxaden, and fluazifop but not clethodim was utilized as the resistant chloroplastic ACCase source compared to known susceptible (S) ACCase. Dose-response studies with sethoxydim, clethodim, fluazifop-p-butyl, and pinoxaden (0.6 to 40 µM) were conducted to compare the ACCase enzyme-herbicides interaction of R1, R2, and S using the malachite green functional assay. Assay results indicated that R biotypes required more ACCase-targeting herbicides to inhibit ACCase activity compared to S. IC50 values of all four herbicides for R biotypes were consistently an order of magnitude greater than S. No sequencing differences in the carboxyltransferase domain was observed for R1 and R2, however, R2 IC50 values were greater across all herbicides. These results indicate the malachite green functional assay is effective in evaluating ACCase enzyme activity of R and S biotypes in the presence of ACCase-targeting herbicides, which can be used as a replacement for the 14C-based radiometric functional assay.

A Strategy for Thioamide Incorporation During Fmoc Solid-Phase Peptide Synthesis with Robust Stereochemical Integrity

2019 ◽  
Author(s):  
luis camacho III ◽  
Bryan J. Lampkin ◽  
Brett VanVeller
Keyword(s):  
Amino Acid ◽  
Functional Groups ◽  
Peptide Synthesis ◽  
Solid Phase ◽  
Solid Phase Peptide Synthesis ◽  
Side Chains ◽  
Amino Acid Side Chains ◽  
Peptide Elongation

We describe a method to protect the sensitive stereochemistry of the thioamide—in analogy to the protection of the functional groups of amino acid side chains—in order to preserve the thioamide moiety during peptide elongation.<br>

The Action of Thrombin on Modified Fibrinogen

1983 ◽  
Vol 49 (03) ◽  
pp. 208-213
Author(s):  
A J Osbahr
Keyword(s):  
Physical Properties ◽  
Negative Charge ◽  
Lysine Residue ◽  
Fibrinopeptide A ◽  
Amino Groups ◽  
Lysine Residues ◽  
Citraconic Anhydride

SummaryThe modification of canine fibrinogen with citraconic anhydride modified the ε-amino groups of the fibrinogen and at the same time generated additional negative charges into the protein. The addition of thrombin to the modified fibrinogen did not induce polymerization; however, the fibrinopeptide was released at a faster rate than from the unmodified fibrinogen. The physical properties of the citraconylated fibrinogen were markedly altered by the modification of 50-60 lysine residues in one hour. A modified fibrinopeptide-A was released by thrombin from the modified fibrinogen and was electrophoretically more anionic than the unmodified fibrinopeptide-A. Edman analysis confirmed the modification of the lysine residue present in the peptide. The rate of removal of citraconylated fibrinopeptide-A from modified fibrinogen by thrombin was 30 to 40 percent greater than the cleavage of unmodified fibrinopeptide-A from unmodified fibrinogen. However, the modification of 60 or more lysine residues in the fibrinogen produced a decrease in the rate of cleavage of citraconylated fibrinopeptide-A. The results suggest that additional negative charge in the vicinity of the attachment of fibrinopeptide-A to canine fibrinogen aids in the removal of the peptide by thrombin.

Sign in / Sign up

Export Citation Format

Share Document