Inna K. Sakodinskaya

Large Activation of Serine Proteases by Pretreatment with Crown Ethers

Pretreatment of serine proteases by lyophilisation in the presence of crown ethers leads to large enhancements of enzyme activity in organic solvents.

Comparative study of the mechanism of alkynelation of ortho-palladated benzylamines and acetanilides

Abstract The ortho -palladated derivate of N,N -dimethylbenzylamine, the complex [Pd(OAc)(C 6 H 4 CH 2 -NMe 2 ) 2 , reacts with styrene in acetic acid solution with a final product of o -Me 2 NCH 2 C 6 CHCHC 6 -H 5 . Two parallel second-order pathways, are involved catalysed, respectively by acetic acid ( k 3 ) and perchloric acid ( k H ). The rate constants k 3 and k H at 30°C are 0.021± 0.005 and 6.8 ± 0.5 dm 3 mol −1 s −1 , while the p K a is 4.6. In the case of the corresponding sacetanilide derivative, the complex [Pd(OAc)(C 6 H 4 NHCOMe)] 2 , the catalysis by perchloric acid is much weaker than for the amine complex, the k 3 and k H being 0.0206 ± 0.0005 and 0.049 ± 0.003 dm 3 mol −1 s −1 , respectively. The p K a in this case is 4.5. Despite the great difference in basicity of N,N -dimethylbenzylamine and acetanilide, the values for p K a are very similar, providing evidence that the perchloric acid-catalysed pathway involves the protonation of groups of comparable basicity in both complexes, that is to say the bridging acetates, while the great variation in the values of k H suggests the existence of other route of acid catalysis for amine complexes. The chloro-bridged acetanilided complex [PdCl(C 6 H 4 NHCOMe)] 2 reacts with styrene in various organic solvents without the perchloride acid-catalysed pathway ( k H ). The Hammett equations log k 3 = −(1.58 ± 0.02) − (1.57 ± 0.06)σ m and log k 3 = −(1.49 ± 0.02) − (0.79 ± 0.15)σ P we obtained in acetic acid solution at 30°C varying ring-substituents in the acetanilide complex and styrene, respectively. On the basis of these and previously reporte results the mechanism of the k 3 pathway for acetanilide complexes is discussed.

Activity and enantioselectivity of serine proteases in transesterification reactions in organic media

The activity and enantioselectivity of α-chymotrypsin and subtilisin Carlsberg in the transesterification of a series of N-acetyl-alanine and -phenylalanine esters in cyclohexane were investigated at constant water activity, both in the absence and presence of 18-crown-6. Isosteric variation of the leaving ability of the alcoholate group in the substrates by fluoro substitution provided information about the acylation step of the reaction. For less reactive esters, the rate of acylation is determined by expulsion of the leaving group, whereas for activated esters the rate is dictated by a physical step, most likely a relatively slow conformational change of the enzyme. This makes the enantioselectivity of the enzymes strongly dependent of the substrate activity and the composition of the reaction medium. The catalytic effect of 18-crown-6, observed for all reactions. can be attributed to an enhanced enzyme-substrate binding and an increase of the rate of acylation.

Initial state and transition state solvation in oxidative dimerization of styrenein the presence of Palladium(II) acetate in various solvents

Abstract Kinetics of oxidative dimerization of styrene to give E,E,-1,4-diphenyl-1,3-butadiene in the presence of palladium(II) acetate has been studied in 12 solvents. In all solvents studied, the reaction obeyed the second order kinetics. Activation parameters (ΔH ≠ , ΔS ≠ , and ΔG ≠ ) were evaluated and isokinetic relationship between ΔH ≠ and ΔS ≠ was established. The trimeric structure of palladium(II) acetate has been supported by spectrophotometric measurements in all solvents except acetonitrile. From the solubility data of palladium(II) acetate its Gibbs free energies of transfer (δμ o Pd ) from heptane (reference solvent) to the respective solvent were calculated. The same values for styrene (δμ o St ) were obtained from the partition constants of styrene between solvents studied and the phase of cross-linked polymer. From δμ o Pd , δμ o St , and δΔ values transfer functions of transition state δμ ≠ were calculated. Correlations of transfer functions of the initial state and the transition state with empirical solvent parameters were examined and used in discussion of a reaction mechanism.

Raman spectroscopy of tris-(hydroxymethyl)aminomethane as a model system for the studies of α-chymotrypsin activation by crown ether in organic solvents

Abstract Raman spectroscopy is employed to study tris-(hydroxymethyl)aminomethane and its complexes with 18-crown-6. The results obtained are used to interpret the known effect of α-chymotrypsin activation by crown ether in organic solvents. Raman spectra of the samples lyophilized from aqueous solutions at various pH values are measured in solid state, acetonitrile and cyclohexane.

Crown ether activation of cross-linked subtilisin Carlsberg crystals in organic solvents

The activity of cross-linked subtilisin Carlsberg crystals in the catalysis of peptide bond formation can be significantly enhanced by pretreatment of the enzyme crystals with crown ethers. Soaking of the enzyme crystals in a solution of crown ether in acetonitrile followed by evaporation of the solvent results in an up to 13 times enhanced enzymatic activity. The effects of crown ether treatment under various conditions gives support for the hypothesis that removal of bound water molecules from the active site during the drying process is the origin of the observed enzyme activation.

Boric acid effect ont he hydrolysis of 4-nitrophenyl 2,3-dihydroxybenzoate: mimic of borate inhibition of serine proteases

Boric acid inhibits the hydrolysis of deprotonated 4-nitrophenyl 2,3-dihydroxybenzoate which proceeds iwth intramolecular general base assistance. This effect is opposite to previously reported borate catalysis of the hydrolysis of salycilic acid esters and mimics borate inhibition of serine proteases.

Kinetics and mechanism of vinylation of ortho-palladated NN-dialkylbenzylamines by para-substituted styrenes

The kinetics of vinylation of ortho-palladated NN-dialkylbenzylamines (1a–g) by CH2CHC6H4R3-p(R3= H, Cl, Br, Me, and MeO) to from the respective 2-dialkylaminomethylstilbenes (2) has been studied mainly at 30°C in acetic acid solvent. The reaction follows second-order kinetics: d[(2)]/dt=k2[(1)][styrene]. The rate constants k2 increase markedly in the presence of both alkali-metal perchlorates (MClO4) and small concentrations of perchloric acid, k2 being directly proportional to [HClO4]. It is shown that the salt effect originates from the solvolytic reaction MClO4+ HOAc ⇌ MOAc + HClO4. Conversion of dimer (1a) into the respective monomer (3) stops the reaction, but the latter species readily forms π-complexes with styrenes. The stability constants of 1 : 1 π-complexes have been determined and their Hammett correlation with a slope of –1.87 is established. For a series of substituted styrenes lgk2 is the linear function of Hammett σp values with a slope ca.–1. For a series of ring-substituted complexes (1) lgk2 correlates with the pKa of the parent benzylamines with a slope of 0.98. The results are interpreted in terms of a mechanism involving intermediate protonation of (1), π-co-ordination of alkene, and subsequent rate-determining intramolecular insertion of styrene into a Pd–C bond through a four-centred transition state.