Epsilon-amino Group Of Lysine Research Articles

On the determination of available lysine in casein and rapeseed protein concentrates using 2,4,6-trinitrobenzenesulphonic acid (TNBS) as a reagent for free epsilon amino group of lysine

The determination of available lysine was studied using 2,4,6-trinitrobenzene-sulfonic acid (TNBS) as a reagent for free epsilon amino group of lysine. By establishing optimal conditions for the hydrolysis of TNP-labelled protein, a modification of a previously described TNBS-method was proposed. The changes reduced the analytical variations substantially from the original method and made it possible to obtain in a simple and rapid way quite reproducible values for the contents of available lysine in casein and rapeseed protein concentrates.

Inhibition of thymidylate synthetase and dihydrofolate reductase by naturally occurring oligoglutamate derivatives of folic acid.

Naturally occurring oligoglutamate derivatives of folic acid in extracts of Escherichia coli have been isolated on the basis of their inhibitory actions toward thymidylate synthetase and dihydrofolate reductase. The inhibitor of thymidylate synthetase has been identified as N-5-formyl-H4pteroyloligoglutamate (approximately 5 amino acid residues). It is 150-fold more inhibitory than the monoglutamate. Synthetic N-5-formyl derivatives containing 3 to 6 glutamyl residues were prepared and found to be 67- to 200-fold more inhibitory than the monoglutamate. N-5-Formimino-H4pteroyltriglutamate is one-twentieth as inhibitory as the corresponding N-5-formyl derivative. The inhibitor of mouse leukemia dihydrofolate reductase has been identified as N-10-formylpteropentaglutamate. It is approximately 7 times as inhibitory as N-10-formylpteroylmonoglutamate. It is 4,400 times as inhibitory toward mouse leukemia dihydrofolate reductase compared with the enzyme from E. coli. Lysine analogs of N-5-formyl-H4folate containing alpha0glutamyl groups in peptide linkage to the epsilon-amino group of lysine were relatively poor inhibitors of thymidylate synthetase. The inhibitory action of folic acid oligoglutamates on E. coli thymidylate synthetase was subject to reversal with 0.4 M NaCl, an effect that was more marked with various pteroyloligoglutamates than with H4homopteroylmonoglutamate and N-5, N-8-deaza-N-10-methylpteroylmonoglutamate.

Trinitrophenylation of the bilirubin binding site of human serum albumin.

Human serum albumin has been modified with 2,4,6-trinitrobenzenesulphonic acid and picryl chloride in low ratios of reagents/albumin. The derivatives have been investigated by spectrophotometry and by thin layer chromatography of the hydrolysates in order to assess the specificity of the reagents. The same reaction conditions were used to modify albumin previously complexed with bilirubin in the ratio of 1:1. The affinity of bilirubin to the modified albumins was estimated by an improved perozidase method. It is concluded that TNBS and picryl chloride react almost quantity with epsilon-amino groups of lysine on the albumin molecule. The results also suggest that at least on TNBS reactive amino group and at least one picryl chloride reactive amino group are located in or near the high-affinity bilirubin binding site.

Identical behavior of the two active sites of myosin with respect to trinitrophenylation.

Myosin and its active subfragments were trinitrophenylated under conditions in which mainly the active site(s) was modified. Proteins modified at the active site(s) could be separated by affinity chromatography on agarose-ATP columns. By two independent methods, ATPase activity measurements and analysis of elution patterns on agarose-ATP columns, it was shown that the introduction of two trinitrophenyl groups per myosin or one per heavy meromyosin subfragment 1 molecule is responsible for the remarkable change in the ATPase activities. Heavy meromyosin subfragment 1 prepared from trinitrophenylated myosin retained the original degree of trinitrophenylation per "active head." The kinetic constant of trinitrophenylation of the epsilon-amino group of lysine at the active site was found to be 2000 S-1-M-1, whereas a much smaller constant of 2.2 S-1-M-1 was obtained for the trinitrophenylation of the unessential lysyl residues of myosin. By using affinity chromatography, we could follow the formation of mono- and ditrinitrophenyl myosin. The amounts of these myosin derivatives at various extents of the reaction corresponded approximately to the calculated amounts, assuming a random and independent trinitrophenylation of the two myosin "heads." It is concluded that in each of the two heads of myosin there is one ATPase active site and these two sites behave in an identical manner with respect to trinitrophenylation.

Effect of carbamylation of lysine epsilon-amino groups on the activity of blood group specific glycoproteins.

Carbamylation of epsilon-amino groups of lysine of human blood group MM glycoprotein, some of its precursors and the blood group A B antigens gave products with 41% - 91% epsilon-amino group substitution. Even the most extensive carbamylation led to only marginal changes in the circular dichroic (CD) spectra of these substances and none in sedimentation coefficients studied. Nevertheless, carbamylation resulted in either increased or unchanged or decreased inhibitory activity of all blood group antigens tested depending solely on the source of the hemagglutinin used. Carbamylation of epsilon-amino groups of these blood group glycoproteins therefore leads to minor conformational changes, not involved with the primary blood group specificity, which is recognized by a large proportion but not by all corresponding antibodies and lectins.

Lactose-amino acid interactions in rations.

Heating of lactose-containing diets was shown to reduce markedly their nutritive value to growing rats; the growth inhibition caused by the inclusion of 5% lactose could be overcome by lysine supplementation. This indicated that the Maillard reaction, involving a reaction between lactose and the epsilon amino group of lysine, was occurring. A model system, using the growing rat fed autoclaved diets, was used to evaluate various feed additives for their potential significance in inducing or inhibiting the Maillard reaction. Alfalfa was found to increase strongly the severity of the Maillard reaction. Ethanol extraction removed some of the active fraction; alfalfa ash and alfalfa saponin were both very potent stimulators of the Maillard reaction. The level and type of added fat was also found to influence the response to lactose in the diet. Ammonium sulphate (1 %) reduced the severity of browning, while urea (1%), copper sulphate (250 ppm), zinc sulphate (100 ppm) and calcium carbonate (4%) had no effect. Glutamic acid (1%), glutamine (0.5%), sodium bicarbonate (5%), sodium propionate (1%) and sodium bisulphite (100 ppm) increased the browning effect.

Selectively blocked and des-histidine-glucagons: preparation and effects on hepatic adenylate cyclase activity.

Recently developed techniques permit selective acylation of the alpha or epsilon amino groups of glucagon. These techniques allow an Edrnan degradation cycle with protection and recovery of the epsilon amino group of lysine. The isolated and purified products were tested for biological activity by measuring stimulation of hepatic glucagon-responsive adenyl cyclase. Both alpha and epsilon amino groups and N-terminal histidine are necessary for biological activity of glucagon (Endocrinology 90: 597, 1972)

Chemical studies on the structure of mucopeptide isolated from Streptococcus bovis.

Mucopeptides isolated from Streptococcus bovis cell walls were found to be composed of alanine, glutamic acid, lysine, and threonine in a mole ratio of 3:1:1:1. A dipeptide, N(epsilon)-lysylthreonine, was isolated from S. bovis mucopeptide by ion-exchange chromatography. This finding suggests that threonine is associated with the bridge which cross-links adjacent tetrapeptides by connecting the epsilon-amino group of lysine of one tetrapeptide to the carboxyl group of d-alanine of another to form the mucopeptide matrix.

Aggregation of platelets by collagen.

In studying some of the properties of collagen responsible for the ability to aggregate platelets it was found that thermal treatment at pH 2.5 of acid-soluble human collagen resulted in a sharp reduction in relative viscosity and platelet aggregating activity at about 35 degrees C. The reduction in viscosity is known to be associated with structural transition from triple helical to random coil form and it is postulated that the native structure of collagen is essential for its platelet aggregation effect. Blockage of the free amino groups by deamination, N-acetylation, or treatment with dinitrofluorobenzene resulted in over 90% reduction in platelet aggregating activity. Addition of cationic proteins to collagen, removal of the negatively charged telopeptides by treatment with pepsin, or acetylation of the free carboxyl groups did not significantly affect the platelet aggregating activity of collagen. On the basis of these findings it is suggested that the free amino groups and specifically the epsilon amino groups of lysine are critical for the platelet aggregating activity of collagen whereas the carboxyl groups are of relatively little importance.

The differential determination of lysine in heated milk. II. Comparison of the in vitro methods with the biological evaluation.

AbstractHaving secured the values for lysine content and availability in a set of milk samples by four in vitro methods, we checked the results with those obtained by the in vivo procedure. The biological determination of lysine availability was performed in young rats using a gluten‐zein basal diet supplemented with the appropriate amino acids. The live‐weight gain was used as a measure of the available lysine content of the samples. The results procured by the biological evaluation correlated best with those of the enzymic determination (ALV‐e) and the corrected FDNB method (ALV‐f II). The absolute figures were identical within the 5% confidence limits. All the data presented are in conformity with the working hypothesis that in heated milk the reaction at the epsilon amino group of lysine controls biological lysine availability by transforming this amino acid into an enzyme‐resistant inactivated form.

The interrelationship of dietary copper and amine oxidase in the formation of elastin

Recent investigations have revealed that copper deficiency results in abnormal elastin formation ( O'Dell et al. , 1961 ; Coulson and Carnes, 1963; Shields et al. , 1962 ; Starcher et al. , 1964 ). The abnormality is characterized by both a decrease in elastin synthesis and an increase in the lysine content of the elastin that is synthesized. Partridge et al. (1964) reported that the cross linkage groups of elastin are synthesized from lysine residues and proposed that the epsilon amino group of lysine was oxidized to an aldehyde prior to condensation to the cross-linkage groups, desmosine and isodesmosine. Such a reaction would be catalyzed by an amine oxidase. The fact that those amine oxidases which have been isolated contain copper ( Yamada and Yasunobu, 1962; Hill and Mann, 1962; Buffoni and Blaschko, 1964) led to the speculation that the role of copper in elastin formation is a reflection of its role in amine oxidase activity. The investigations presented in this report were undertaken to examine this hypothesis.

INVESTIGATION OF THE ACTIVE SITES OF LYSOZYME. CARBOXYMETHYLATION OF THE IMIDAZOLE GROUP OF HISTIDINE AND OF THE EPSILON-AMINOGROUP OF LYSINE.

INVESTIGATION OF THE ACTIVE SITES OF LYSOZYME. CARBOXYMETHYLATION OF THE IMIDAZOLE GROUP OF HISTIDINE AND OF THE EPSILON-AMINOGROUP OF LYSINE.

Contribution of E-amino groups to ninhydrin color production in proteins.

The ninhydrin color given by model compounds and by characterized proteins shows a consistent contribution from the epsilon-amino group of lysine of about 67 percent of that of an alpha-amino group, except in free lysine or in N-terminal lysine, where the epsilon-amino group makes a small contribution (7 to 10 percent) to the total color. This information can be applied to structure determination of N(epsilon) peptides of lysine.