Jack bean urease (EC 3.5.1.5). V. On the mechanism of action of urease on urea, formamide, acetamide, N-methylurea, and related compounds

1980 ◽  
Vol 58 (12) ◽  
pp. 1335-1344 ◽  
Author(s):  
Nicholas E. Dixon ◽  
Peter W. Riddles ◽  
Carlo Gazzola ◽  
Robert L. Blakeley ◽  
Burt Zerner
Keyword(s):  
Enzymatic Hydrolysis ◽  
Mechanism Of Action ◽  
Active Site ◽  
Ph Dependence ◽  
Jack Bean Urease ◽  
Nickel Ion ◽  
Jack Bean ◽  
First Time ◽  
Hydrolysis Of ◽  
Hydrolysis Of Urea

Acetamide and N-methylurea have been shown for the first time to be substrates for jack bean urease. In the enzymatic hydrolysis of urea, formamide, acetamide, and N-methylurea at pH 7.0 and 38 °C, kcat has the values 5870, 85, 0.55, and 0.075 s−1, respectively. The urease-catalyzed hydrolysis of all these substrates involves the active-site nickel ion(s). Enzymatic hydrolysis of the following compounds could not be detected: phenyl formate, p-nitroformanilide, trifluoroacetamide, p-nitrophenyl carbamate, thiourea, and O-methylisouronium ion. In the enzymatic hydrolysis of urea, the pH dependence of kcat between pH 3.4 and 7.8 indicates that at least two prototropic forms are active. Enzymatic hydrolysis of urea in the presence of methanol gave no detectable methyl carbamate. A mechanism of action for urease is proposed which involves initially an O-bonded complex between urea and an active-site Ni2+ ion and subsequently an O-bonded carbamato–enzyme intermediate.

Jack bean urease (EC 3.5.1.5). III. The involvement of active-site nickel ion in inhibition by β-mercaptoethanol, phosphoramidate, and fluoride

1980 ◽  
Vol 58 (6) ◽  
pp. 481-488 ◽  
Author(s):  
Nicholas E. Dixon ◽  
Robert L. Blakeley ◽  
Burt Zerner
Keyword(s):  
Dissociation Constant ◽  
Active Site ◽  
Competitive Inhibitor ◽  
Jack Bean Urease ◽  
Nickel Ion ◽  
Nickel Ions ◽  
Jack Bean ◽  
Complex Fluoride ◽  
Spectral Changes ◽  
Hydrolysis Of

Interaction of β-mercaptoethanol with urease produces large, rapid and fully reversible spectral changes in that part of the electronic absorption spectrum which is associated with the tightly bound nickel ions. The spectrophotometrically determined value of the dissociation constant of the β-mercaptoethanol–urease complex (0.95 ± 0.05 mM at pH 7.12 and 25 °C) is in agreement with the Ki (0.72 ± 0.26 mM) for β-mercaptoethanol acting as a competitive inhibitor in the hydrolysis of urea. This constitutes direct evidence that the nickel in jack bean urease is at the active site. Inhibition of urease by phosphoramidate is slowly achieved and slowly reversed, and upon reactivation of the isolated phosphoramidate–urease complex, phosphoramidate is regenerated in good yield. Spectrophotometric experiments indicate that phosphoramidate binds to nickel ion in urease. Competition with β-mercaptoethanol was used to determine a dissociation constant (1.23 ± 0.10 mM at pH 7.12 and 25 °C) for a fluoride–urease complex in which fluoride ion also coordinates with an active-site nickel ion. Kinetic evidence is presented which indicates that in the presence of urea, a ternary complex (fluoride–urea–urease) is formed.

New Method for the Synthesis of a Mononucleating Cyclic Peptide Ligand, Crystal Structures of its Ni, Zn, Cu, and Co Complexes, and Their Inhibitory Bioactivity Against Urease

2007 ◽  
Vol 60 (5) ◽  
pp. 375 ◽  
Author(s):  
Kui Cheng ◽  
Zhong-Lu You ◽  
Hai-Liang Zhu
Keyword(s):  
Cyclic Peptide ◽  
Peptide Ligand ◽  
Jack Bean Urease ◽  
One Pot ◽  
Nickel Ion ◽  
Jack Bean ◽  
Metal Free ◽  
Synthetic Procedure ◽  
First Time ◽  
Solvothermal Conditions

A novel cyclic peptide complex, NiL 1 (H2L = 12,24-dihydroxy-1,6-dioxo-2,5,14,17-tetraaza[6*6]metacyclophane-13,17-diene has been synthesized for the first time under solvothermal conditions through a one-pot synthetic procedure using nickel ion as the template reagent. It was found that other metal ions were not suitable for the direct template reagent in this reaction. The nickel ion was eliminated from the complex and the metal-free cyclic peptide ligand H2L was obtained through a series of reactions. Then, ZnII, CuII, and CoII were coordinated with H2L under the same solvothermal conditions to produce three isomorphous complexes ZnL 2, CuL 3, and CoL 4. Their inhibitory bioactivities against urease were then studied. The copper(ii) complex 3 was the strongest inhibitor against jack bean urease, while H2L, 2, and 4 showed weak or no inhibitory activity against this enzyme.

Jack bean urease (EC 3.5.1.5). IV. The molecular size and the mechanism of inhibition by hydroxamic acids. Spectrophotometric titration of enzymes with reversible inhibitors

1980 ◽  
Vol 58 (12) ◽  
pp. 1323-1334 ◽  
Author(s):  
Nicholas E. Dixon ◽  
John A. Hinds ◽  
Ann K. Fihelly ◽  
Carlo Gazzola ◽  
Donald J. Winzor ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Active Site ◽  
Binding Sites ◽  
Spectrophotometric Titration ◽  
Hydroxamic Acids ◽  
Jack Bean Urease ◽  
Equivalent Weight ◽  
Guanidinium Chloride ◽  
Jack Bean ◽  
Equilibrium Ultracentrifugation

Kinetic, spectral, and other studies establish that hydroxamic acids bind reversibly to active-site nickel ion in jack bean urease. Equilibrium ultracentrifugation studies establish that the molecular weight of native urease is 590 000 ± 30 000 while that of the subunit formed in 6 M guanidinium chloride in the presence of β-mercaptoethanol is ~95 000. Essentially the same subunit molecular weight (~93 000) is found by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, subsequent to denaturation in a guanidinium chloride – β-mercaptoethanol system at various temperatures. Coupled with an equivalent weight of 96 600 for binding of the inhibitors acetohydroxamic acid and phosphoramidate, these results establish securely that urease is a hexamer with one active site per 96 600-dalton subunit. Consistent values for the equivalent weight are obtained by a routine spectrophotometric titration of the active site of freshly prepared urease with trans-cinnamoylhydroxamic acid. General equations are derived which describe spectrophotometric titrations of binding sites of any enzyme with a reversible inhibitor. These equations allow the evaluation of the difference spectrum of the protein–inhibitor complex even when the binding sites cannot readily be saturated with the inhibitor or vice versa.

Synthesis, cytotoxic and urease inhibitory activities of some novel isatin-derived bis-Schiff bases and their copper(ii) complexes

MedChemComm ◽  
2016 ◽  
Vol 7 (5) ◽  
pp. 914-923 ◽  
Author(s):  
Humayun Pervez ◽  
Maqbool Ahmad ◽  
Sumera Zaib ◽  
Muhammad Yaqub ◽  
Muhammad Moazzam Naseer ◽  
...  
Keyword(s):  
Schiff Bases ◽  
Active Compound ◽  
Active Site ◽  
Binding Mode ◽  
Jack Bean Urease ◽  
Jack Bean ◽  
Inhibitory Activities ◽  
Urease Inhibitory

The putative binding mode of the most active compound 3b in the active site of Jack bean urease.

Enzymatic Hydrolysis of Carotenoid Fatty Acid Esters of Red Pepper (Capsicum annuum L.) by a Lipase from Candida rugosa

2000 ◽  
Vol 55 (11-12) ◽  
pp. 971-975 ◽  
Author(s):  
Dietmar Ernst Breithaupt
Keyword(s):  
Enzymatic Hydrolysis ◽  
Candida Rugosa ◽  
Fatty Acid Esters ◽  
Red Pepper ◽  
Free Form ◽  
Enzymatic Cleavage ◽  
Reaction Conditions ◽  
First Time ◽  
Hydrolysis Of ◽  
Acid Esters

Analyses of red pepper extracts which had been pretreated with lipase type VII (EC 3.1.1.3.) from Candida rugosa showed for the first time pepper carotenoid esters to be substrates of this enzyme. However, the extent of enzymatic hydrolysis depends on the respective carotenoid and was not quantitative compared to chemical saponification. After enzymatic cleavage, 67-89% of total capsanthin, 61-65% of total zeaxanthin, 70-81% of total β-cryptoxanthin and 70-86% of total violaxanthin were detected in free form. Nevertheless, the method described here offers the possibility to cleave in part several carotenoid esters originating from red pepper quickly and under comparatively mild reaction conditions. Replacement of the generally performed alkaline hydrolysis by enzymatic cleavage allows the resulting product to be used in food industry as “natural” coloring agent e.g. to colour cheese and jellies.

Monolithic gelation of chitosan solutions via enzymatic hydrolysis of urea

2006 ◽  
Vol 64 (3) ◽  
pp. 419-424 ◽  
Author(s):  
A. Chenite ◽  
S. Gori ◽  
M. Shive ◽  
E. Desrosiers ◽  
M.D. Buschmann
Keyword(s):  
Enzymatic Hydrolysis ◽  
Hydrolysis Of ◽  
Hydrolysis Of Urea

scholarly journals The nickel ion environment in jack bean urease

1984 ◽  
Vol 220 (2) ◽  
pp. 591-595 ◽  
Author(s):  
L Alagna ◽  
S S Hasnain ◽  
B Piggott ◽  
D J Williams
Keyword(s):  
Fine Structure ◽  
Structure Study ◽  
Jack Bean Urease ◽  
Model Compounds ◽  
Deprotonated Form ◽  
Nickel Ion ◽  
Edge Structure ◽  
Jack Bean ◽  
X Ray ◽  
X Ray Absorption

Preliminary results of an extended X-ray absorption fine structure (e.x.a.f.s.) and X-ray absorption near edge structure study of jack bean urease have recently been reported [Hasnain & Piggott (1983) Biochem. Biophys. Res. Commun. 112, 279]. These results indicate that the environment of the nickel ion in the enzyme is similar to that in the model compounds Ni(L)2(L')1(ClO4)1 (where L is 1-n-propyl-2-alpha-hydroxybenzylbenzimidazole and L' is the deprotonated form) and Ni(HMB)3(Br)2 (where HMB is 2-hydroxymethylbenzimidazole), the closest similarity being with Ni(L)2-(L')1(ClO4)1. A detailed e.x.a.f.s. analysis has now been carried out and the crystal structures of the two model compounds solved. These results are reported here.

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