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The glycoprotein nature of pig kidney diamine oxidase. Role of disulphide groups and arginine residues in the concanavalin A-diamine oxidase interaction.

ABSTRACT: Pig kidney diamine oxidase (DAO) was found to contain 5% (w/w) natural hexose, 3.25% glucosamine, 2.61% N-acetylglucosamine and 0.25% N-acetylneuraminic acid. The enzyme exhibited strong affinity towards concanavalin A (Con A) with a stoichiometry of 1:4.6. The kinetics of interaction approached an apparent first-order rate, with a rate constant (Kapp.) value of 1.5 x 10(-2) min-1. The enzyme reduced with dithiothreitol followed by alkylation with iodoacetamide showed an increase in the stoichiometry of the Con A-DAO interaction. Similarly arginine modification by phenylglyoxal caused decreased affinity, with an altered Kapp. value of 9.09 x 10(-3) min-1. The results suggest that, besides the carbohydrate content, the protein moiety of the enzyme also plays a significant role in the Con A-DAO interaction.

SUBMITTER: Shah MA

PROVIDER: S-EPMC1149263 | biostudies-other | 1988 Jul

REPOSITORIES: biostudies-other

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Inhibition of pig kidney diamine oxidase by substrate analogues.

Project description:1. The oxidation of p-dimethylaminomethylbenzylamine was followed spectrophotometrically by measuring the change in E(250) caused by the p-dimethylaminomethylbenzaldehyde produced. 2. This reaction was inhibited by substrate analogues such as isothiouronium, guanidino, dimethylsulphonium and trimethylammonium compounds. 3. The inhibition by both mono- and bis-onium compounds has been studied and a comprehensive theory is developed to explain both the type and degree of inhibition produced.

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The dynamic disulphide relay of quiescin sulphydryl oxidase.

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Structure and inhibition of human diamine oxidase.

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Evidence for the importance of arginine residues in pig kidney alkaline phosphatase.

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| S-EPMC1161134 | biostudies-other

A new crystal form of human diamine oxidase.

Project description:Copper amine oxidases (CAOs) are ubiquitous in nature and catalyse the oxidative deamination of primary amines to the corresponding aldehydes. Humans have three viable CAO genes (AOC1-3). AOC1 encodes human diamine oxidase (hDAO), which is the frontline enzyme for histamine metabolism. hDAO is unique among CAOs in that it has a distinct substrate preference for diamines. The structure of hDAO in space group P2(1)2(1)2(1) with two molecules in the asymmetric unit has recently been reported. Here, the structure of hDAO refined to 2.1 A resolution in space group C222(1) with one molecule in the asymmetric unit is reported.

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Oxidation of p-dimethylaminomethylbenzylamine by pig kidney diamine oxidase. A new method for spectrophotometric assay.

Project description:1. The oxidation of some para-substituted benzylamines by diamine oxidase produces the corresponding aldehydes. This was studied to develop a spectrophotometric method for following the enzyme reaction, as the aldehydes produced absorb strongly at 250nm where the substrates are almost optically transparent. 2. p-Dimethylaminomethylbenzylamine was the most useful substrate and full details of its preparation are given. The synthesis of its related oxidation product, p-dimethylaminomethylbenzaldehyde, is also described. 3. The effects of variations in pH, ionic strength, temperature and oxygenation on the reaction are described and the usefulness of the method is illustrated by several applications and assessed by comparison with the standard spectrophotometric assay.

| S-EPMC1178797 | biostudies-other

Dynamic nature of disulphide bond formation catalysts revealed by crystal structures of DsbB.

Project description:In the Escherichia coli system catalysing oxidative protein folding, disulphide bonds are generated by the cooperation of DsbB and ubiquinone and transferred to substrate proteins through DsbA. The structures solved so far for different forms of DsbB lack the Cys104-Cys130 initial-state disulphide that is directly donated to DsbA. Here, we report the 3.4 A crystal structure of a DsbB-Fab complex, in which DsbB has this principal disulphide. Its comparison with the updated structure of the DsbB-DsbA complex as well as with the recently reported NMR structure of a DsbB variant having the rearranged Cys41-Cys130 disulphide illuminated conformational transitions of DsbB induced by the binding and release of DsbA. Mutational studies revealed that the membrane-parallel short alpha-helix of DsbB has a key function in physiological electron flow, presumably by controlling the positioning of the Cys130-containing loop. These findings demonstrate that DsbB has developed the elaborate conformational dynamism to oxidize DsbA for continuous protein disulphide bond formation in the cell.

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Interaction of two arginine residues in lactate oxidase with the enzyme flavin: conversion of FMN to 8-formyl-FMN.

Project description:Two arginine residues, Arg-181 and Arg-268, are conserved throughout the known family of FMN-containing enzymes that catalyze the oxidation of alpha-hydroxyacids. In the lactate oxidase from Aerococcus viridans, these residues have been changed to lysine in two single mutations and in a double mutant form. In addition, Arg-181 has been replaced by methionine to determine the effect of removing the positive charge on the residue. The effects of these replacements on the kinetic and thermodynamic properties are reported. With all mutant forms, there are only small effects on the reactivity of the reduced flavin with oxygen. On the other hand, the efficiency of reduction of the oxidized flavin by l-lactate is greatly reduced, particularly with the R268K mutant forms. The results demonstrate the importance of the two arginine residues in the binding of substrate and its interaction with the flavin, and are consistent with a previous hypothesis that they also play a role of charge neutralization in the transition state of substrate dehydrogenation. The replacement of Arg-268 by lysine also results in a slow conversion of the 8-CH(3)- substituent of FMN to yield 8-formyl-FMN, still tightly bound to the enzyme, and with significantly different physical and chemical properties from those of the FMN-enzyme.

| S-EPMC27174 | biostudies-literature

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