scholarly journals The acetylation of insulin

1971 ◽  
Vol 121 (5) ◽  
pp. 737-745 ◽  
Author(s):  
D. G. Lindsay ◽  
S. Shall
Keyword(s):  
Biological Activity ◽  
Free Amino ◽  
Tertiary Structure ◽  
Group Analysis ◽  
Insulin Antibodies ◽  
The Other ◽  
Amino Group ◽  
Amino Groups ◽  
Chloromethyl Ketone ◽  
End Group

The acetylation of the free amino groups of insulin was studied by reaction of the hormone with N-hydroxysuccinimide acetate at pH6.9 and 8.5. The products formed were separated by chromatography on DEAE-Sephadex and were characterized by isoelectric focusing, by end-group analysis, by the incorporation of [3H]acetyl groups in the molecule, and by treatment with trypsin that had been treated with 1-chloro-4-phenyl-3-toluene-p-sulphonamidobutan-2-one (‘tosylphenylalanyl chloromethyl ketone’). Three monosubstituted products, two disubstituted products and one trisubstituted derivative were prepared. The α-amino groups of the terminal residues and the ∈-amino group of the lysine-B29 were the sites of reaction. Acetylation of any of the free amino groups did not affect the biological activity of insulin. It was demonstrated, however, that substitution at the glycine-A1 amino group by the larger residues, acetoacetyl or thiazolidinecarbonyl, produced a decrease in biological activity. Modification of the lysine-B29 or phenylalanine-B1 amino groups with these larger reagents did not affect the biological activity. Modification of the phenylalanine-B1 amino group by any of the three substituents resulted in a large decrease in the affinity of insulin for anti-insulin antibodies raised in the guinea pig. Modification of the other two amino groups did not affect the reaction with antibody. These observations are correlated with the tertiary structure of insulin.

scholarly journals IMMUNOLOGIC PRODUCTION OF ANTIANGIOTENSIN

1960 ◽  
Vol 111 (3) ◽  
pp. 419-427 ◽  
Author(s):  
Sharad D. Deodhar
Keyword(s):  
Biological Activity ◽  
Angiotensin Ii ◽  
Aromatic Amines ◽  
Free Amino ◽  
Parent Compound ◽  
The Other ◽  
Amino Group ◽  
Angiotensin I ◽  
Absolute Requirement ◽  
Specific Activities

Angiotensin II was coupled with bovine γ-globulin (BGG) through the following series of reactions. See PDF for Structure By determinations of the aromatic amine and tyrosine contents of p-aminobenzoylangiotensin II, the number of p-aminobenzoyl residues introduced per molecule of angiotensin II was calculated. Absorption spectra (between 250 and 500 mµ) of BGG complexes of p-aminobenzoylangiotensin II and six different para substituted aromatic amines were compared. Specific activities (dog units/millimicromole) of the different intermediate products were determined. Presence of a terminal, free amino group does not appear to be an absolute requirement for the biological activity of angiotensin II, since substitution of a p-aminobenzoyl radical in this group yields a product with 40 to 50 per cent of the activity of the parent compound. Angiotensin I, on the other hand, is completely inactivated under identical circumstances. Possible implication of this finding has been discussed.

scholarly journals The reversible reaction of protein amino groups with exo-cis-3,6-endoxo-Δ4-tetrahydrophthalic anhydride. The reaction with lysozyme

1970 ◽  
Vol 118 (5) ◽  
pp. 733-739 ◽  
Author(s):  
M. Riley ◽  
R. N. Perham
Keyword(s):  
Free Amino ◽  
Tertiary Structure ◽  
Complete Recovery ◽  
Enzymic Activity ◽  
Carboxyl Group ◽  
Amino Group ◽  
Half Time ◽  
Amino Groups ◽  
Model Compounds ◽  
Anchimeric Assistance

1. The reaction of exo-cis-3,6-endoxo-Δ4-tetrahydrophthalic anhydride with amino groups of model compounds and lysozyme is described. 2. Reaction with the ∈-amino group of Nα-acetyl-l-lysine amide gives rise to two diastereoisomeric products; at acid pH the free amino group is liberated with anchimeric assistance by the neighbouring protonated carboxyl group with a half-time of 4–5h at pH3.0 and 25°C. 3. The amino groups of lysozyme can be completely blocked, with total loss of enzymic activity. Dialysis at pH3.0 results in complete recovery of the native primary and tertiary structure of lysozyme and complete return of catalytic activity. 4. The specificity of reaction of this and other anhydrides with amino groups in proteins is discussed.

scholarly journals Acetoacetylation of insulin

1969 ◽  
Vol 115 (3) ◽  
pp. 587-595 ◽  
Author(s):  
D. G. Lindsay ◽  
S. Shall
Keyword(s):  
Tertiary Structure ◽  
Group Analysis ◽  
Amino Group ◽  
Hormonal Activity ◽  
Fourth Component ◽  
End Group

Insulin was treated with diketen at pH6·9. The reaction mixture was resolved into four components by DEAE-Sephadex chromatography. The first component was unchanged insulin. The second and third components were shown by end-group analysis to be substituted on phenylalanine B-1 and glycine A-1 respectively. The fourth component was disubstituted on both phenylalanine B-1 and glycine A-1. The ∈-amino group of lysine B-29 was not involved in the reaction at low reagent concentrations. The purity of these derivatives was checked by their electrophoretic behaviour and by measurement of the rate of their reaction with trinitrobenzenesulphonic acid. The hormonal activity of the derivatives was determined. The effect of the modifications on the hormonal activity and the tertiary structure of insulin is discussed.

COMPARATIVE STUDIES OF THE POTENTIATION OF CORTICOTROPHIN ACTIVITY BY SEVERAL INACTIVE DERIVATIVES

1963 ◽  
Vol 42 (2) ◽  
pp. 209-213 ◽  
Author(s):  
Arthur I. Cohen ◽  
Edward H. Frieden
Keyword(s):  
Biological Activity ◽  
Comparative Studies ◽  
Free Amino ◽  
Hormonal Activity ◽  
Amino Groups

ABSTRACT A number of corticotrophin analogues have been prepared, some of which potentiate the biological activity of the untreated hormone in vitro. The free amino groups of corticotrophin appear to be essential not only for hormonal activity, but also for the interaction of the analogues with the tissue corticotrophin inactivating system which is assumed to account for the potentiating effect.

Effect of cross-linking agents on insulin associated responses in adipocytes

1982 ◽  
Vol 60 (10) ◽  
pp. 987-1000 ◽  
Author(s):  
H. Joseph Goren ◽  
C. Ronald Kahn
Keyword(s):  
Insulin Receptor ◽  
Glucose Transport ◽  
Free Amino ◽  
Glucose Oxidation ◽  
Insulin Binding ◽  
Cross Linking ◽  
Amino Group ◽  
Amino Groups ◽  
Disuccinimidyl Suberate ◽  
Reactive Groups

The effect of 10 bifunctional cross-linking agents and four monofunctional analogues was studied on isolated adipocytes. [125I]Insulin binding and degradation, basal and insulin-stimulated glucose oxidation, and 3-O-methyl glucose uptake were measured. Two cross-linkers, which possess succinimide ester residues (disuccinimidyl suberate and dithiobis(succinimidyl propionate)) and react selectively with amino groups, appeared to react relatively specifically with the insulin receptor. Both produced a slight stimulation of basal glucose transport and metabolism, a marked inhibition of insulin-stimulated glucose transport and metabolism, and a marked decrease in insulin binding. Pretreatment of cells with unlabelled insulin partially blocked the effect of disuccinimidyl suberate, and as has been previously shown, disuccinimidyl suberate cross-linked insulin to its receptor. A monofunctional analogue of these compounds was 100-fold less active in altering cellular metabolic activity. Bisimidates, such as dimethyl suberimidate, dimethyl adipimidate, and dimethyl dithiobispropionimidate, also react with free amino groups but are more hydrophilic. These agents produced similar effects on glucose oxidation as the succinimide esters, but had little or no effect on insulin binding. The effects of these agents are not blocked by insulin and they do not cross-link insulin to its receptor. Mixed bifunctional reagents containing either a succinimide ester or an imidate and a group which reacts with thiols produced effects similar to the cross-linkers containing two succinimide groups or bisimidates, respectively. The bifunctional arylating agents difluorodinitrobenzene and bis(fluoronitrophenyl)sulfone produce marked effects on insulin binding and glucose oxidation at micromolar concentrations, but the monofunctional analogue fluorodinitrobenzene is almost equally active suggesting that with these compounds chemical modifications and not cross-linking was important. With neither the mixed bifunctional reagents, nor the arylating agents, did insulin pretreatment alter the effect of cross-linker and none of these agents cross-linked [125I]insulin to its receptor. These data suggest that the insulin receptor possesses a free amino group in a hydrophobic environment in its active site. A reactive amino group in a hydrophilic environment as well as other reactive groups are also present in some component of the insulin receptor–effector complex. Chemical modification or cross-linking of these functional groups results in an inhibition or mimicking of insulin action. Further study will be required to identify the exact locus of these sites.

New Method for the Determination of Free Amino Groups in Intact Pure Proteins: Relationship to Available Lysine

1976 ◽  
Vol 59 (6) ◽  
pp. 1251-1254
Author(s):  
James M Purcell ◽  
Daniel J Quimby ◽  
James R Cavanaugh
Keyword(s):  
Bovine Serum Albumin ◽  
Bovine Serum ◽  
Free Amino ◽  
Amino Group ◽  
Amino Groups ◽  
Primary Amino ◽  
Available Lysine ◽  
Β Lactoglobulin ◽  
Group Concentration

Abstract A new rapid method for the quantitative and routine determination of free amino groups in intact pure proteins has been developed. Primary amino groups are labeled with fluorescamine and the labeled groups are detected by absorption spectroscopy in the range 375–390 nm. The amino group concentration can be determined in a few minutes without hydrolyzing the labeled protein and extracting a lysine derivative. The method was tested with the following proteins: lysozyme, α-lactalbumin, β-lactoglobulin, bovine serum albumin, ribonuclease, ribonuclease-S-peptide, and αsl-rasciii B. Application of this method to the estimation of available lysine is discussed.

Modified Hydroxyl Value Assay and Application in Analysis of Molecular Ingredient of Polylactic Acid Oligomers

2011 ◽  
Vol 306-307 ◽  
pp. 692-695
Author(s):  
Zhi Ping Qiu ◽  
Ji Da Chen ◽  
Juan Bu ◽  
Rong De Shu
Keyword(s):  
Fourier Transform ◽  
Infrared Spectroscopy ◽  
Fourier Transform Infrared Spectroscopy ◽  
Polylactic Acid ◽  
1H Nmr ◽  
Group Analysis ◽  
The Other ◽  
Hydroxyl Value ◽  
Modified Method ◽  
End Group

A modification was proposed to overcome problems of inaccuracy while traditional end group analysis was applied for hydroxyl value assay of polylactic acid oligomer owing to hydrolyzation. The modified method possessed simplification and accuracy resulting from avoiding hydrolyzation of polylactic acid oligomers, and it could be used for characterising molecular ingredient of polylactic acid oligomers, while the other techniques, such as 1H-NMR, Fourier transform infrared spectroscopy and viscosimetry, were applied at the same time.

A Study of the Conformational Properties of Glucagon in the Presence of Glycols

1974 ◽  
Vol 52 (6) ◽  
pp. 456-468 ◽  
Author(s):  
C. C. Contaxis ◽  
R. M. Epand
Keyword(s):  
Biological Activity ◽  
Tertiary Structure ◽  
Structural Response ◽  
Spectroscopic Properties ◽  
The Other ◽  
Monomeric Form ◽  
Conformational Properties ◽  
Α Helix

Elimination of the polymerization of glucagon has been achieved in the presence of glycols. The monomeric form of glucagon, found to be exclusively present under these conditions, is studied for its structural response to changes in the hydrophobicity of the environment. A reversible folding of the α-helix from about 0 to 80% is found to occur and it is suggested to proceed in two sequentially formed helical segments. Changes in the other spectroscopic properties are interpreted as changes in the tertiary structure. Implications for the biological activity are discussed.

scholarly journals The reaction of 2,4,6-trinitrobenzenesulphonic acid with amino acids, peptides and proteins

1968 ◽  
Vol 108 (3) ◽  
pp. 383-391 ◽  
Author(s):  
R. B. Freedman ◽  
G. K. Radda
Keyword(s):  
Amino Acids ◽  
Glutamate Dehydrogenase ◽  
Free Amino ◽  
Reactive Species ◽  
Second Order ◽  
Amino Group ◽  
Exponential Rate ◽  
Amino Groups ◽  
Sh Group ◽  
Kinetics Of

1. The kinetics of the reaction of 2,4,6-trinitrobenzenesulphonic acid with various amino acids, peptides and proteins were studied by spectrophotometry. 2. The reaction of the α- and ∈-amino groups in simple amino acids was found to be second-order, and the unprotonated amino group was shown to be the reactive species. 3. By allowing for the concentration of unreactive −NH3+ group, intrinsic reactivities for the free amino groups were derived and shown to be correlated with the basicities. 4. The SH group of N-acetylcysteine was found to be more reactive to 2,4,6-trinitrobenzenesulphonic acid than most amino groups. 5. The reactions of insulin, chymotrypsinogen and ribonuclease with 2,4,6-trinitrobenzenesulphonic acid were analysed in terms of three exponential rate curves, each referring to one or more amino groups of the proteins. 6. The reaction of lysozyme with 2,4,6-trinitrobenzenesulphonic acid was found to display an acceleration effect. 7. From the reaction of 2,4,6-trinitrobenzenesulphonic acid with glutamate dehydrogenase at several enzyme concentrations, it was possible to discern two sets of amino groups of different reactivity, and to show that the number of groups in each set was decreased by aggregation of the enzyme.

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