Statistical Calculations of the Accuracy of the Michaelis Constant from Viscosimetric Determinations of Polymetaphosphatase and Dextranase Activity.

Hultin, Eskil; Bohman, Gerd; Santesson, Johan; Holmberg, Pär; Eriksson, G.; Blinc, R.; Paušak, S.; Ehrenberg, L.; Dumanović, J.

doi:10.3891/acta.chem.scand.21-1575

Cited by 14 publications

(6 citation statements)

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“…Initial velocity values were defined by the change in the viscosity min-' (mg of dextranase)-'. The inverse values of the specific viscosities were plotted versus time for each digestion experiment according to Hultin [27], and straight lines were obtained. This indicates that the dextranase hydrolyses all or almost all of the glycosidic linkages with equal ease and that it is an endo-dextranase.…”

Section: Lineweaver-burk Plots Of the Digestion Experiments Using Ementioning

confidence: 99%

Dextran-Linked Purine Nucleosides as Substrates and Inhibitors of Adenosine Deaminase

1982

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Dextran-bound adenosine, inosine, and nebularine have been prepared by carbodiimide coupling of their 2',3'-O-(4-carboxyethyl-l-methylbutylidene) cyclic acetal derivatives to 6-aminohexyldextran or 12-aminododecanyldextran. The latter polymers were prepared by cyanogen-bromide activation of dextran T80 followed by reaction with 1,6-diaminohexane or 1 ,12-diaminododecane. A high CNBr concentration leads to high-molecular-weight material, probably due to cross-linking, accompanied by a decrease in the digestion velocity using endo-dextranase from Penicillium species (EC 3.2.1.1 1).The dextran-bound nucleosides, as well as the nucleoside 2', 3'-O-(4-ethoxycarbonyI-l -methylbutylidene) acetal derivatives, were tested as substrates and inhibitors for adenosine deaminase. The K, of the adenosine acetal ester is identical to that of adenosine which shows that acetalation does not hinder complex formation. Since the maximum velocity of deamination is decreased fourfold, the modified substrate does not fit as well as the nucleoside. The polymer-bound acetals show a 3 -8-fold increase of K, or Ki and unchanged V compared to the corresponding acetals while dextranase digestion of the support does not alter the kinetic data. This indicates that the length of the polysaccharide chain does not interfere either with the complex formation or with the catalytic activity of the modified substrate. Since the activation energies of the deamination reactions of adenosine, its acetal ester, and dextran-linked adenosine are all similar (29.8 -32.3 kJ mol-') it is concluded that no diffusion control of the enzymatic reaction results from the binding of the nucleoside acetals to dextran T80.Heterogeneous enzyme-catalysed reactions can be divided into two categories: (a) reactions between soluble substrates and enzymes embedded in biological membranes or other native complex structures ; (b) reactions between a soluble enzyme and a polymer-bound or insoluble substrate [l -61.This prompted us to immobilise substrates and inhibitors of adenosine deaminase via cyclic acetal derivatives onto soluble dextran T80. Dextrans are a group of structurally related polysaccharides (mr, lo5 -10') elaborated by various microorganisms, especially by strains of Leuconostoc. All dextrans contain linear chains of 1 .+ 6-linked a-D-glucopyranose residues as the dominant structural feature [7]. Since we have already immobilised adenosine deaminase on dextran T80 [8], we now have a complete kinetic model system in which either the enzyme or its binding partners are covalently bound to a polymer matrix.The polymer attachment of nucleosides and nucleoside antimetabolites has several implications. (a) From the pharmacological point of view, the brief plasma persistance of small substrates and inhibitors can be overcome by linking the biomolecule to a polymer such as dextran T80, which is used as a blood volume expander. High activity of the modified substrate can only be expected if the position of attachment is carefully chosen and, if possible, a stereochemica...

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Section: Lineweaver-burk Plots Of the Digestion Experiments Using Ementioning

confidence: 99%

Dextran-Linked Purine Nucleosides as Substrates and Inhibitors of Adenosine Deaminase

1982

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“…There are abundant sources of dextranase, which can be produced by higher organisms and microorganisms. Since the first report of Cellvibrio fulva dextranase in 1940 (Hultin et al, 1967), more than 100 patents have been issued on dextran-hydrolyzing enzymes found in a number of microbial groups (Abdelwahed et al, 2014;Deng et al, 2020;Dong et al, 2021;Huang et al, 2019;Khalikova et al, 2003;Lee et al, 2010;H. Liu et al, 2019;N.…”

Section: Dextranasementioning

confidence: 99%

Research progress on structure and catalytic mechanism of dextranase

Xia

et al. 2023

eFood

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Dextranase is an important industrial enzyme for the preparation of isomaltooligosaccharides. According to the crystal structures of 20 dextranases of four classes, we summarized the structural characteristics of enzymes and the binding modes of enzymes and ligands in this paper. Based on the characteristics of the products, we analyzed the characteristics of the binding domain and functions of dextranase by means of molecular simulation and molecular docking. The relationship between the product specificity of dextranase, its catalytic pocket shape, and its catalytic mode was briefly summarized. The catalytic mechanism of dextranase was systematically discussed, which provided the basis and reference for the rational transformation of dextranase.

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“…The kinetic constants K M and V MAX of the purified enzyme were calculated by fitting the PG activity data determined in a 0.09-9 g/L PGA concentration range to a linear regression after applying the Hanes-Hultin equation [Hultin, 1967]. The inhibition constant, K i , for the end products of PGA degradation was calculated after applying the Dixon equation [Dixon, 1953] to the PG activity data measured with increasing amounts of the end products, at 0.375 and 0.4375 g/L PGA.…”

Section: Determination Of Kinetic Parametersmentioning

confidence: 99%

Purification and Biochemical and Kinetic Properties of an Endo-Polygalacturonase from the Industrial Fungus <b><i>Aspergillus sojae</i></b>

Fratebianchi

Cavello

Cavalitto³

2017

Microb Physiol

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An endo-polygalacturonase secreted by Aspergillus sojae was characterized after being purified to homogeneity from submerged cultures with orange peel as the sole carbon source by gel filtration and ion-exchange chromatographies. According to SDS-PAGE and analytical isoelectric focusing analyses, the enzyme presents a molecular weight of 47 kDa and pI value of 4.2. This enzyme exhibits considerable stability under highly acidic to neutral conditions (pH 1.5-6.5) and presents a half-life of 2 h at 50°C. Besides its activity towards pectin and polygalacturonic acid, the enzyme displays pectin-releasing activity, acting best in a pH range of 3.3-5.0. Thin-layer chromatographic analysis revealed that tri-galacturonate is the main enzymatic end product of polygalacturonic acid hydrolysis, indicating that it is an endo-polygalacturonase. The enzyme exhibits Michaelis-Menten kinetics, with K_M and V_MAX values of 0.134 mg/mL and 9.6 µmol/mg/min, respectively, and remained stable and active in the presence of SO₂, ethanol, and various cations assayed except Hg²⁺.

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Statistical Calculations of the Accuracy of the Michaelis Constant from Viscosimetric Determinations of Polymetaphosphatase and Dextranase Activity.

Cited by 14 publications

References 0 publications

Dextran-Linked Purine Nucleosides as Substrates and Inhibitors of Adenosine Deaminase

Dextran-Linked Purine Nucleosides as Substrates and Inhibitors of Adenosine Deaminase

Research progress on structure and catalytic mechanism of dextranase

Purification and Biochemical and Kinetic Properties of an Endo-Polygalacturonase from the Industrial Fungus <b><i>Aspergillus sojae</i></b>

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