Role Of Histidine Residues Research Articles

Proton nuclear magnetic resonance study on the roles of histidine residues in the binding of polypeptide chain elongation factor Tu from Thermus thermophilus with aminoacyl transfer ribonucleic acid and guanine nucleotides.

Proton nuclear magnetic resonance (1H NMR) spectra were measured of the polypeptide chain elongation factor Tu (EF-Tu) from an extreme thermophile, Thermus thermophilus HB8 [Nakano, A., Miyazawa, T., Nakamura, S., & Kaziro, Y. (1979) Arch. Biochem. Biophys. 196, 233-238], in order to elucidate the environment around functionally important histidine residues. In the present study, the behavior of five histidine C2 proton signals was studied in more detail. A hydrogen-deuterium exchange experiment was carried out on the histidine C2 protons of free EF-Tu, and the previous assignments of C2 proton signals were revised in part. An analysis of the 1H NMR spectra of EF-Tu photooxidized under various conditions indicates that a histidine residue is located in the aminoacyl-tRNA binding site and is probably essential for the binding with aminoacyl-tRNA. A solvent-accessible histidine residue is found to lie near the aminoacyl-tRNA binding site. Furthermore, the effect of paramagnetic hexacyanochromate(III) ion on the 1H NMR spectra of free EF-Tu suggests that another histidine residue lies near the guanine nucleotide binding site.

Delta aminolaevulinate dehydratase: its mechanism of action

1. 1. Taking into account available experimental data. a mechanism of action for aminolaevulinate dehydratase is proposed. 2. 2. Besides the formation of a Schiff base between the substrate and the enzyme, the existence of a minimal functional dimer necessary for activity is also considered, as well as certain characteristics of the type of interactions involved in the inter-subunit contact sites and the role of histidine residues in the transport of protons from and to the active center.

Mechanism of an active transport of calcium. Ethoxyformylation of sarcoplasmic reticulum vesicles.

Ethoxyformylation of sarcoplasmic reticulum vesicles is performed to study a possible role of histidine residues in the calcium translocation process. The influence of the chemical modification is evaluated on the Ca2+-dependent ATPase activity, and on the Ca2+ uptake parameters: VCa (initial rate of calcium uptake) and CCa (amount of cation accumulated at the steady state). The substitution of the amino acids is monitored by three different techniques: (a) by amino acid analysis of the ethoxyformylated material further submitted to modification by diazonium-1-H-tetrazole, or by sulfhydryl titration using 5-5'-dithiobis (2-nitrobenzoic acid); (b) by 14C labeling followed by the removing of labels after NH2OH or imidazole treatment at pH 7; (c) by spectrophotometric measurements at 230 nm. The ethoxyformylation reaction is not specific for histidine at pH 6.1 and 10 degrees. About 1 lysyl group/mol of ATPase is first modified. Then 1 (with a pseudo-first order rate constant of 240 (+/- 20) 10(-3) min-1) or 2 histidines are modified. No substitution of tyrosine or sulfhydryl groups can be detected under our experimental conditions. A decrease of the Ca2+-dependent ATPase activity correlated with the inhibition of both VCa and Cca corresponds to the chemical substitution of the histidine. No direct correlation between the decrease of the activities and the modification of the lysine can be found. After removing the ethoxyformyl group from the histidine, only the Ca2+-dependent ATPase activity is restored to its initial value. No protection is found when the reaction is performed in the presence of ATP or p-nitrophenylphosphate. These results can be explained if one assumes that the ethoxyformylation of the histidine residue(s) induces a conformational change modifying the affinity of the membrane for nucleotides.

Essential Role of Histidine Residues for the Catalytic Activity of Ribose Phosphate Isomerase

Essential Role of Histidine Residues for the Catalytic Activity of Ribose Phosphate Isomerase

Evidence for an essential histidine in carboxypeptidase Y. Reaction with the chloromethyl ketone derivative of benzyloxycarbonyl-L-phenylalanine.

The possible role of histidine residues in the catalytic function of carboxypeptidase Y from bakers' yeast has been investigated using site-specific reagents. Among the reagents tested, benzyloxy-L-phenylalanylchloromethane (Z-PheCH2Cl) was the most powerful inhibitor of the enzyme. It irreversibly inactivated both the peptidase and esterase activities with an apparent second order rate constant of 3.8 M-minus 1 S-minus 1; the D isomer caused essentially no effect on either activity. Inhibition by L-Z-PheCH2Cl, the reaction retarded by certain competitive inhibitors of the enzyme. Using radioactive L-Z-PheCH2Cl, the reaction with the enzyme was shown to be essentially stoichiometric. Diisopropylphosphorofluoridate (iPr2PF)-inactivated enzyme failed to react with Z-PheCH2Cl, and conversely, the Z-PheCH2Cl-inhibited enzyme failed to react with radioactive iPr2PF. Amino acid analyses of the Z-PheCH2Cl-inactivated enzyme revealed the loss of essentially 1 residue, with a concomitant yield of a 0.62 residue of N-t-carboxymethylhistidine. Since carboxypeptidase Y has a reactive serine at its active center, we concluded from these results that the mechanism involves a charge-relay system in the hydrolysis of peptide and ester substrates, as in chymotrypsin. An -SH group of carboxypeptidase Y was not affected during the reaction with L-Z-PheCH2Cl. The generic name "serine carboxypeptidase" has been proposed for carboxypeptidase Y and for the iPr2PF-sensitive carboxypeptidases from plants, molds, and animal tissues, in order to distinguish them from "metal carboxypeptidase" to which carboxypeptidase A (EC 3.4.12.2) and B (EC 3.4.12.3) belong.

The role of histidine residues in yeast hexokinase. Enzymic and structural studies.

Specific acylation of the histidine residues of yeast hexokinase with diethylpyrocarbonate at pH 6.1 or 7.5 leads to a partial and reversible inactivation of the enzyme.The carboethoxylation of the histidine residues at pH 7.5 was faster than at 6.1 and the loss in activity was also higher.However the difference in the amount of inactivation was related to a greater instability of the carboethoxylated enzyme at pH 7.5.When the carboethoxylation was carried out at pH 7.5 and at 0 °C, only the histidines residues were modified (nine residues per enzyme subunit) and still 40% of the activity was left. d‐Glucose protects the enzyme activity in the course of the carboethoxylation at pH 7.5, but has no effect at pH 6.1. However the rate and number of histidines acylated were the same as when the experiments were carried in absence of the substrate.All these results clearly indicate that the histidine residues are not implicated either in the catalytic or in the substrate binding site of yeast hexokinase.This was also confirmed by showing that the 60% inactivated enzyme had the same Km as the native enzyme for both substrates d‐glucose and Mg · ATP. Only v was affected.The inactivation effect of the carboethoxylation was not related to a displacement of the pH optimum of the activity, since the enzymes species having five and eight histidine residues modified‐respectively in absence and presence of d‐glucose at pH 7.5 gave the same pH profile of activity as the native enzyme.Absorption spectroscopy, fluorescence and ultracentrifugation studies have shown that the loss of enzymic activity in the course of the modification of the histidine residues is only related to changes of the protein conformation.

The Role of Histidine Residues of Yeast Alcohol Dehydrogenase

The Role of Histidine Residues of Yeast Alcohol Dehydrogenase

The role of histidine residues in glutamate dehydrogenase

1. Glutamate dehydrogenase was subject to rapid inactivation when irradiated in the presence of Rose Bengal or incubated in the presence of ethoxyformic anhydride. 2. Inactivation in the presence of Rose Bengal led to the photo-oxidation of four histidine residues. Oxidation of three histidine residues had little effect on enzyme activity, but oxidation of the fourth residue led to the almost total loss of activity. 3. Acylation of glutamate dehydrogenase with ethoxyformic anhydride at pH6.1 led to the modification of three histidine residues with a corresponding loss of half the original activity. Acylation at pH7.5 led to the modification of two histidine residues and a total loss of enzyme activity. 4. One of the histidine residues undergoing reaction at pH6.1 also undergoes reaction at pH7.5. 5. The presence of either glutamate or NAD(+) in the reaction mixtures at pH6.1 had no appreciable effect. At pH7.5 glutamate caused a marked decrease in both the degree of alkylation and degree of inactivation. NAD(+) had no effect on the degree of inactivation at pH7.5 but did modify the extent of acylation. 6. The normal response of the enzyme towards ADP was unaffected by acylation at pH6.1 or 7.5. 7. The normal response of the enzyme towards GTP was altered by treatment at both pH6.1 and 7.5.

Inhibition of acylneuraminate pyruvate-lyase: evidence of intermediary Schiff's base formation and of a possible role of histidine residues.

Inhibition of acylneuraminate pyruvate-lyase: evidence of intermediary Schiff's base formation and of a possible role of histidine residues.