Mechanism of serine hydroxymethylase catalyzed cleavage of L-erythro-.beta.-phenylserine: pH dependence of elementary kinetic processes from spectroscopic, pre-steady-state kinetic, and competitive inhibition studies

Biochemistry ◽  
1979 ◽  
Vol 18 (5) ◽  
pp. 821-830 ◽  
Author(s):  
Wei-Mei Ching ◽  
Roland G. Kallen
Keyword(s):  
Steady State ◽  
Competitive Inhibition ◽  
Ph Dependence ◽  
Steady State Kinetic ◽  
Kinetic Processes ◽  
Inhibition Studies ◽  
State Kinetic

scholarly journals A steady-state kinetic study of the reaction catalysed by the secondary-amine mono-oxygenase of Pseudomonas aminovorans

1976 ◽  
Vol 157 (1) ◽  
pp. 197-205 ◽  
Author(s):  
D F Brook ◽  
P J Large
Keyword(s):  
Steady State ◽  
Affinity Chromatography ◽  
Secondary Amine ◽  
Gel Filtration ◽  
Product Inhibition ◽  
Steady State Kinetic ◽  
Ping Pong ◽  
Michaelis Constants ◽  
Inhibition Studies ◽  
State Kinetic

1. Secondary-amine mono-oxygenase (proposed EC group 1.14.99.-) was partially purified from trimethylamine-grown Pseudomonas aminovorans by (NH4)2SO4 fractionation, gel filtration, hydrophobic chromatography on 5-aminopentylamino-Sepharose, and affinity chromatography on Sepharose-bound NADH. 2. Some problems in the affinity-chromatography step are discussed. 3. A steady-state kinetic analysis varying substrate, oxygen and electron-donor concentrations was performed, which, over the concentration range studied, gave a series of families of approximately parallel double-reciprocal plots. From secondary and tertiary plots, Michaelis constants of 0.160 mM, 0.086 mM and 0.121 mM were obtained for dimethylamine, NADPH and oxygen respectively. 4. Product-inhibition studies supported the postulated Hexa Uni Ping Pong (triple-transfer) reaction mechanism.

scholarly journals Variation in the pH-dependent pre-steady-state and steady-state kinetic characteristics of cysteine-proteinase mechanism: evidence for electrostatic modulation of catalytic-site function by the neighbouring carboxylate anion

2003 ◽  
Vol 372 (3) ◽  
pp. 735-746 ◽  
Author(s):  
Syeed HUSSAIN ◽  
Surapong PINITGLANG ◽  
Tamara S. F. BAILEY ◽  
James D. REID ◽  
Michael A. NOBLE ◽  
...  
Keyword(s):  
Steady State ◽  
Cysteine Proteinase ◽  
Ph Dependence ◽  
Catalytic Site ◽  
Rate Determining Step ◽  
Steady State Kinetic ◽  
Pka Value ◽  
Site Function ◽  
Hydrolysis Of ◽  
State Kinetic

The acylation and deacylation stages of the hydrolysis of N-acetyl-Phe-Gly methyl thionoester catalysed by papain and actinidin were investigated by stopped-flow spectral analysis. Differences in the forms of pH-dependence of the steady-state and pre-steady-state kinetic parameters support the hypothesis that, whereas for papain, in accord with the traditional view, the rate-determining step is the base-catalysed reaction of the acyl-enzyme intermediate with water, for actinidin it is a post-acylation conformational change required to permit release of the alcohol product and its replacement in the catalytic site by the key water molecule. Possible assignments of the kinetically influential pKa values, guided by the results of modelling, including electrostatic-potential calculations, and of the mechanistic roles of the ionizing groups, are discussed. It is concluded that Asp161 is the source of a key electrostatic modulator (pKa 5.0±0.1) in actinidin, analogous to Asp158 in papain, whose influence is not detected kinetically; it is always in the ‘on’ state because of its low pKa value (2.8±0.06).

scholarly journals Bulgecin A: a novel inhibitor of binuclear metallo-β-lactamases

2005 ◽  
Vol 387 (3) ◽  
pp. 585-590 ◽  
Author(s):  
Alan M. SIMM ◽  
E. Joel LOVERIDGE ◽  
John CROSBY ◽  
Matthew B. AVISON ◽  
Timothy R. WALSH ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Steady State ◽  
Stenotrophomonas Maltophilia ◽  
Competitive Inhibition ◽  
Zinc Ion ◽  
Glycosidic Linkage ◽  
Sugar Moiety ◽  
Aeromonas Veronii ◽  
Steady State Kinetic ◽  
State Kinetic

Bulgecin A, a sulphonated N-acetyl-D-glucosamine unit linked to a 4-hydroxy-5-hydroxymethylproline ring by a β-glycosidic linkage, is a novel type of inhibitor for binuclear metallo-β-lactamases. Using steady-state kinetic analysis with nitrocefin as the β-lactam substrate, bulgecin A competitively inhibited the metallo-β-lactamase BceII from Bacillus cereus in its two-zinc form, but failed to inhibit when the enzyme was in the single-zinc form. The competitive inhibition was restored by restoring the second zinc ion. The single-zinc metallo-β-lactamase from Aeromonas veronii bv. sobria, ImiS, was not inhibited by bulgecin A. The tetrameric L1 metallo-β-lactamase from Stenotrophomonas maltophilia was subject to partial non-competitive inhibition, which is consistent with a kinetic model in which the enzyme bound to inhibitor retains catalytic activity. Docking experiments support the conclusion that bulgecin A co-ordinates to the zinc II site in metallo-β-lactamases via the terminal sulphonate group on the sugar moiety.

Human liver aldehyde reductase: pH dependence of steady-state kinetic parameters

1991 ◽  
Vol 287 (2) ◽  
pp. 329-336 ◽  
Author(s):  
Aruni Bhatnagar ◽  
Ballabh Das ◽  
Si-Qi Liu ◽  
Satish K. Srivastava
Keyword(s):  
Steady State ◽  
Kinetic Parameters ◽  
Human Liver ◽  
Ph Dependence ◽  
Aldehyde Reductase ◽  
Steady State Kinetic ◽  
State Kinetic

scholarly journals Steady-state kinetic analysis of soluble methane mono-oxygenase from Methylococcus capsulatus (Bath)

1986 ◽  
Vol 236 (1) ◽  
pp. 155-162 ◽  
Author(s):  
J Green ◽  
H Dalton
Keyword(s):  
Steady State ◽  
Kinetic Analysis ◽  
Product Inhibition ◽  
Methylococcus Capsulatus ◽  
Reaction Sequence ◽  
Oxidation Of Methane ◽  
Steady State Kinetic ◽  
Substitution Mechanism ◽  
Inhibition Studies ◽  
State Kinetic

A steady-state kinetic analysis of purified soluble methane mono-oxygenase of Methylococcus capsulatus (Bath) was performed. The enzyme was found to follow a concerted-substitution mechanism. Methane binds to the enzyme followed by NADH, which reacts to yield reduced enzyme and NAD+. The reduced enzyme-methane complex binds O2 to give a second ternary complex, which breaks down to release water and methanol. In this way the enzyme can control the supply of electrons to the active site to coincide with the arrival of methane. Product-inhibition studies (with propylene as substrate) supported the reaction mechanism proposed. Ki values for NAD+ and propylene oxide are reported. The Km for NADH varied from 25 microM to 300 microM, depending on the nature of the hydrocarbon substrate, and thus supports the proposed reaction sequence. With methane as substrate the Km values for methane, NADH and O2 were shown to be 3 microM, 55.8 microM and 16.8 microM respectively. With propylene as substrate the Km values for propylene, NADH and O2 were 0.94 microM, 25.2 microM and 12.7-15.9 microM respectively. Methane mono-oxygenase was shown to be well adapted to the oxidation of methane compared with other straight-chain alkanes.

Steady-State Kinetic and Inhibition Studies of the Mammalian Target of Rapamycin (mTOR) Kinase Domain and mTOR Complexes

Biochemistry ◽  
2010 ◽  
Vol 49 (39) ◽  
pp. 8488-8498 ◽  
Author(s):  
Zhihua Tao ◽  
John Barker ◽  
Stone D.-H. Shi ◽  
Michael Gehring ◽  
Shaoxian Sun
Keyword(s):  
Steady State ◽  
Mammalian Target Of Rapamycin ◽  
Kinase Domain ◽  
Target Of Rapamycin ◽  
Mtor Kinase ◽  
Steady State Kinetic ◽  
Inhibition Studies ◽  
State Kinetic
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