scholarly journals Inverse Solvent Isotope Effects in Enzyme-Catalyzed Reactions

Molecules ◽  
2020 ◽  
Vol 25 (8) ◽  
pp. 1933
Author(s):  
Patrick L. Fernandez ◽  
Andrew S. Murkin
Keyword(s):  
Isotope Effect ◽  
Isotope Effects ◽  
Solvent Isotope Effect ◽  
Kinetic Isotope ◽  
Rate Limiting Step ◽  
Catalyzed Reactions ◽  
Enzyme Catalyzed Reactions ◽  
Solvent Isotope ◽  
Rate Limiting ◽  
Solvent Isotope Effects

Solvent isotope effects have long been used as a mechanistic tool for determining enzyme mechanisms. Most commonly, macroscopic rate constants such as kcat and kcat/Km are found to decrease when the reaction is performed in D2O for a variety of reasons including the transfer of protons. Under certain circumstances, these constants are found to increase, in what is termed an inverse solvent kinetic isotope effect (SKIE), which can be a diagnostic mechanistic feature. Generally, these phenomena can be attributed to an inverse solvent equilibrium isotope effect on a rapid equilibrium preceding the rate-limiting step(s). This review surveys inverse SKIEs in enzyme-catalyzed reactions by assessing their underlying origins in common mechanistic themes. Case studies for each category are presented, and the mechanistic implications are put into context. It is hoped that readers may find the illustrative examples valuable in planning and interpreting solvent isotope effect experiments.

Kinetic and solvent isotope effects in oxidation of halogen derivatives of tyramine catalyzed by monoamine oxidase A

2019 ◽  
Vol 167 (1) ◽  
pp. 49-54
Author(s):  
Małgorzata Pająk
Keyword(s):  
Monoamine Oxidase ◽  
Isotope Effect ◽  
Isotope Effects ◽  
Monoamine Oxidase A ◽  
Oxidative Deamination ◽  
Solvent Isotope Effect ◽  
Kinetic Isotope ◽  
Solvent Isotope ◽  
Derivatives Of ◽  
Solvent Isotope Effects

Abstract The isotope effects approach was used to elucidate the mechanism of oxidative deamination of 3′-halotyramines, catalyzed by monoamine oxidase A (EC 1.4.3.4). The numerical values of kinetic isotope effect (KIE) and solvent isotope effect (SIE) were established using a non-competitive spectrophotometric technique. Based upon KIE and SIE values, some of the mechanistic details of investigated reaction were discussed.

Solvent isotope effects in the oxidation of dipeptides by aqueous chlorine

1999 ◽  
Vol 77 (5-6) ◽  
pp. 997-1004 ◽  
Author(s):  
X L Armesto ◽  
M Canle L. ◽  
V García ◽  
J A Santaballa
Keyword(s):  
Isotope Effect ◽  
Deuterium Oxide ◽  
Isotope Effects ◽  
Second Step ◽  
Solvent Isotope Effect ◽  
General Base ◽  
Peptide Oxidation ◽  
Solvent Isotope ◽  
Hydrolysis Of ◽  
Solvent Isotope Effects

A kinetic study of the mechanism of oxidation of Ala-Gly and Pro-Gly by aqueous chlorine has been carried out. Among other experimental facts, the deuterium solvent isotope effects were used to clarify the mechanisms involved. In a first stage, N-chlorination takes place, and then the (N-Cl)-dipeptide decomposes through two possible mechanisms, depending on the acidity of the medium. The initial chlorination step shows a small isotope effect. In alkaline medium, two consecutive processes take place: first, the general base-catalyzed formation of an azomethine (β ca. 0.27), which has an inverse deuterium solvent isotope effect (kOH-/kOD- ~ 0.8). In a second step, the hydrolysis of the azomethine intermediate takes place, which is also general base-catalyzed, without deuterium solvent isotope effect, the corresponding uncatalyzed process having a normal deuterium solvent isotope effect (kH2O/kD2O ~ 2). In acid medium, the (N-Cl)-dipeptide undergoes disproportionation to a (N,N)-di-Cl-dipeptide, the very fast decomposition of the latter in deuterium oxide preventing a reliable estimation of the solvent isotope effect.Key words: chlorination, deuterium isotope effects, fractionation factors, peptide oxidation, water treatment.

Kinetic Solvent Isotope Effect for the Spontaneous Solvolysis Acetic and Propionic Anhydrides

1971 ◽  
Vol 49 (22) ◽  
pp. 3665-3670 ◽  
Author(s):  
R. E. Robertson ◽  
B. Rossall ◽  
W. A. Redmond
Keyword(s):  
Acetic Anhydride ◽  
Isotope Effect ◽  
Isotope Effects ◽  
Temperature Dependency ◽  
Solvent Isotope Effect ◽  
Propionic Anhydride ◽  
Solvent Isotope ◽  
Hydrolysis Of ◽  
Solvent Isotope Effects

The large kinetic solvent isotope effects for the neutral hydrolysis of acetic and propionic anhydride show unusual temperature dependency; the former passing through a maximum at about 15°, the latter showing a minimum at 30°. This unusual temperature dependency is the consequence of widely different values of the apparent ΔCp≠ in H2O and D2O: the value for acetic anhydride in H2O being −74 ± 2 cal deg−1 mol−1 but −32 ± 4 in D2O. The corresponding values for propionic anhydride being −31 ± 2 in H2O but −94 ± 10 in D2O. The implications of these differences are discussed.

Studies in Solvolysis. Part IV. Substituent and Solvent Isotope Effects in the Solvolysis of a Series of Benzyl Trifluoroacetates

1972 ◽  
Vol 50 (12) ◽  
pp. 1886-1890 ◽  
Author(s):  
June G. Winter ◽  
J. M. W. Scott
Keyword(s):  
Isotope Effect ◽  
Deuterium Oxide ◽  
Isotope Effects ◽  
The Other ◽  
Solvent Isotope Effect ◽  
Other Hand ◽  
Solvent Isotope ◽  
Hydrolysis Of ◽  
Solvent Isotope Effects

The rates of neutral hydrolysis of a series of 4-substituted benzyl trifluoroacetates 4-X-C6H4CH2OCOCF3, X = NO2, Cl, H, CH3, and OCH3 have been studied in water and deuterium oxide, both solvents containing 0.012 mol fraction of acetone. The alteration of the rates with the nature of the 4-substituent and the kinetic solvent isotope effect (k(H2O)/k(D2O)) are consistent with the proposal that the esters with X = NO2, Cl, H, and CH3 all react by an acyl–oxygen BAc2 mechanism. On the other hand, the same mechanistic criteria indicate that the 4-methoxybenzyl ester reacts by both the BAc2 and the SN1 alkyl–oxygen fission paths in equal amounts.

scholarly journals Substrate-Dependent Sensitivity of SIRT1 to Nicotinamide Inhibition

Biomolecules ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 312
Author(s):  
Stacia Rymarchyk ◽  
Wenjia Kang ◽  
Yana Cen
Keyword(s):  
Proton Transfer ◽  
Isotope Effects ◽  
Negative Regulator ◽  
Base Exchange ◽  
Glucose And Lipid Metabolism ◽  
Rate Limiting Step ◽  
Limiting Step ◽  
Catalyzed Reactions ◽  
Solvent Isotope ◽  
Rate Limiting

SIRT1 is the most extensively studied human sirtuin with a broad spectrum of endogenous targets. It has been implicated in the regulation of a myriad of cellular events, such as gene transcription, mitochondria biogenesis, insulin secretion as well as glucose and lipid metabolism. From a mechanistic perspective, nicotinamide (NAM), a byproduct of a sirtuin-catalyzed reaction, reverses a reaction intermediate to regenerate NAD+ through “base exchange”, leading to the inhibition of the forward deacetylation. NAM has been suggested as a universal sirtuin negative regulator. Sirtuins have evolved different strategies in response to NAM regulation. Here, we report the detailed kinetic analysis of SIRT1-catalyzed reactions using endogenous substrate-based synthetic peptides. A novel substrate-dependent sensitivity of SIRT1 to NAM inhibition was observed. Additionally, SIRT1 demonstrated pH-dependent deacetylation with normal solvent isotope effects (SIEs), consistent with proton transfer in the rate-limiting step. Base exchange, in contrast, was insensitive to pH changes with no apparent SIEs, indicative of lack of proton transfer in the rate-limiting step. Consequently, NAM inhibition was attenuated at a high pH in proteated buffers. Our study provides new evidence for “activation by de-repression” as an effective sirtuin activation strategy.

scholarly journals A solvent-isotope-effect study of proton transfer during catalysis by Escherichia coli (lacZ) β-galactosidase

1990 ◽  
Vol 268 (2) ◽  
pp. 317-323 ◽  
Author(s):  
T Selwood ◽  
M L Sinnott
Keyword(s):  
Escherichia Coli ◽  
Isotope Effect ◽  
Kinetic Isotope Effect ◽  
Isotope Effects ◽  
Effect Study ◽  
Solvent Isotope Effect ◽  
Kinetic Isotope ◽  
Solvent Isotope ◽  
Paper Abstract ◽  
Hydrolysis Of

1. Michaelis-Menten parameters for the hydrolysis of 4-nitrophenyl β-D-galactopyranoside and 3,4-dinitrophenyl β-D-galactopyranoside Escherichia coli (lacZ) β-galactosidase were measured as a function of pH or pD (pL) in both 1H2O and 2H2O. 2. For hydrolysis of 4-nitrophenyl β-D-galactopyranoside by Mg2(+)-free enzyme, V is pL-independent below pL 9, but the V/Km-pL profile is sigmoid, the pK values shifting from 7.6 +/- 0.1 in 1H2O to 8.2 +/- 0.1 in 2H2O, and solvent kinetic isotope effects are negligible, in accord with the proposal [Sinnott, Withers & Viratelle (1978) Biochem. J. 175, 539-546] that glycone-aglycone fission without acid catalysis governs both V and V/Km. 3. V for hydrolysis of 4-nitrophenyl β-D-galactopyranoside by Mg2(+)-enzyme varies sigmoidally with pL, the pK value shifting from 9.19 +/- 0.09 to 9.70 +/- 0.07; V/Km shows both a low-pL fall, probably due to competition between Mg2+ and protons [Tenu, Viratelle, Garnier & Yon (1971) Eur. J. Biochem. 20, 363-370], and a high-pL fall, governed by a pK that shifts from 8.33 +/- 0.08 to 8.83 +/- 0.08. There is a negligible solvent kinetic isotope effect on V/Km, but one of 1.7 on V, which a linear proton inventory shows to arise from one transferred proton. 4. The variation of V and V/Km with pL is sigmoid for hydrolysis of 3,4-dinitrophenyl β-D-galactopyranoside by Mg2(+)-enzyme, with pK values showing small shifts, from 8.78 +/- 0.09 to 8.65 +/- 0.08 and from 8.7 +/- 0.1 to 8.9 +/- 0.1 respectively. There is no solvent isotope effect on V or V/Km for 3,4-dinitrophenyl β-D-galactopyranoside, despite hydrolysis of the galactosyl-enzyme intermediate governing V. 5. Identification of the ‘conformation change’ in the hydrolysis of aryl galactosides proposed by Sinnott & Souchard [(1973) Biochem. J. 133, 89-98] with the protolysis of the magnesium phenoxide arising from the action of enzyme-bound Mg2+ as an electrophilic catalyst rationalizes these data and also resolves the conflict between the proposals and the 18O kinetic-isotope-effect data reported by Rosenberg & Kirsch [(1981) Biochemistry 20, 3189-3196]. It should be noted that the actual Km values were determined to higher precision than can be estimated from the Figures in this paper.(ABSTRACT TRUNCATED AT 400 WORDS)

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