Simple conformation space search protocols for the evaluation of enantioselectivity of lipases

Orrenius, Christian; Heusden, C. Van; Ruiten, Jippe Van; Overbeeke, P.L.Antoine; Kierkels, Hans; Duine, J. A.; Jongejan, Jaap A.

doi:10.1093/protein/11.12.1147

Cited by 15 publications

(8 citation statements)

References 2 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Whether this causes structural changes or leaves an empty space within the enzyme remains unknown until the structure is solved. The fact that Δ R‐S Δ H ‡ significantly increased for W104H would result in an erroneous prediction of a very high enantioselectivity if only enthalpic energy considerations were made, as is done in most contemporary molecular modeling of enzyme catalysis where the entropic components generally are neglected (Hæffner et al 1998; Ke et al 1998; Orrenius et al 1998a,b). Attempts to calculate activation entropy in enzyme catalysis with computer modeling have recently appeared in literature (Strajbl et al 2000; Villa et al 2000).…”

Section: Discussionmentioning

confidence: 99%

Rational design of enantioselective enzymes requires considerations of entropy

et al. 2001

View full text Add to dashboard Cite

Entropy was shown to play an equally important role as enthalpy for how enantioselectivity changes when redesigning an enzyme. By studying the temperature dependence of the enantiomeric ratio E of an enantioselective enzyme, its differential activation enthalpy (⌬ R-S ⌬H ‡ ) and entropy (⌬ R-S ⌬S ‡ ) components can be determined. This was done for the resolution of 3-methyl-2-butanol catalyzed by Candida antarctica lipase B and five variants with one or two point mutations. ⌬ R-S ⌬S ‡ was in all cases equally significant as ⌬ R-S ⌬H ‡ to E. One variant, T103G, displayed an increase in E, the others a decrease. The altered enantioselectivities of the variants were all related to simultaneous changes in ⌬ R-S ⌬H ‡ and ⌬ R-S ⌬S ‡ . Although the changes in ⌬ R-S ⌬H ‡ and ⌬ R-S ⌬S ‡ were of a compensatory nature the compensation was not perfect, thereby allowing modifications of E. Both the W104H and the T103G variants displayed larger ⌬ R-S ⌬H ‡ than wild type but exhibited a decrease or increase, respectively, in E due to their different relative increase in ⌬ R-S ⌬S ‡ .

show abstract

Section: Discussionmentioning

confidence: 99%

Rational design of enantioselective enzymes requires considerations of entropy

et al. 2001

View full text Add to dashboard Cite

show abstract

“…In the past, there have been many other attempts to evaluate enantiomeric ratio by molecular modeling. For instance, enantioselectivity has been correlated, with more or less success, to the difference in the potential energy part ∆U of the free energy difference ∆∆G ‡ between the two enantiomer transition states [11,12,13]. One of the obvious limitations of this kind of calculation is that, while the potential energy of a protein in explicit solvent is typically of the order of several thousand kilocalories per mole, the energy difference between R and S tetrahedral intermediates, which are good models for the transition states [14], is expected to be less than five kilocalories per mole, as the difference in free energy ∆∆G ‡ is related to the enantioselectivity, expressed by the enantiomeric ratio E, as follows:∆∆G ‡ = −RT lnE [15,16,17].…”

Section: Introductionmentioning

confidence: 99%

Contribution of both catalytic constant and Michaelis constant to CALB enantioselectivity: Use of FEP calculations for prediction studies

Chaput

Sanejouand

Balloumi

et al. 2012

Journal of Molecular Catalysis B: Enzymatic

View full text Add to dashboard Cite

Candida antarctica lipase B (CALB) is characterised by its stability and ease of production and is widely used in the pharmaceutical industry. Here we report on the enantioselectivity of the enzyme using both experimental and computational methods. The apparent kinetic parameters were first experimentally determined for enantiopure butan-2-ol and pentan-2-ol substrates. We demonstrate that enantiopreference for the R form of butan-2-ol arises mainly from a lower apparent K M . This corresponds to a major contribution of ∆∆G ES , the free energy difference between the ES complex formed with the R and S enantiomers, to ∆∆G ‡ , the free energy difference between both transition states, in comparison with ∆∆G kcat , the activation free energy difference. In the case of pentan-2-ol, we show that the enantiopreference for the R form comes from both a lower K M and a higher k cat . In addition, we used, for the first time, the Free Energy Perturbation method to evaluate the free energy difference between tetrahedral intermediates formed with R and S alcohol enantiomers for a series of secondary alcohols . This is a valid model for ∆∆G ‡ . Computational results were found to be in good agreement with experimental data, and enable the determination of substrate orientation in the active site with fair confidence.

show abstract

“…Attempts to calculate enantioselectivity using force-field methods have been made~DeTar, 1981; Bemis et al, 1992;Faber et al, 1994;Orrenius et al, 1998!, for example, calculations of energy Fig. 1.…”

mentioning

confidence: 99%

Stereoselectivity of Pseudomonas cepacia lipase toward secondary alcohols: A quantitative model

2000

View full text Add to dashboard Cite

The lipase from Pseudomonas cepacia represents a widely applied catalyst for highly enantioselective resolution of chiral secondary alcohols. While its stereopreference is determined predominantly by the substrate structure, stereoselectivity depends on atomic details of interactions between substrate and lipase. Thirty secondary alcohols with published E values using P. cepacia lipase in hydrolysis or esterification reactions were selected, and models of their octanoic acid esters were docked to the open conformation of P. cepacia lipase. The two enantiomers of 27 substrates bound preferentially in either of two binding modes: the fast-reacting enantiomer in a productive mode and the slow-reacting enantiomer in a nonproductive mode. Nonproductive mode of fast-reacting enantiomers was prohibited by repulsive interactions. For the slow-reacting enantiomers in the productive binding mode, the substrate pushes the active site histidine away from its proper orientation, and the distance d~H NE Ϫ O alc ! between the histidine side chain and the alcohol oxygen increases. d~H NE Ϫ O alc ! was correlated to experimentally observed enantioselectivity: in substrates for which P. cepacia lipase has high enantioselectivity~E Ͼ 100!, d~H NE Ϫ O alc ! is Ͼ2.2 Å for slow-reacting enantiomers, thus preventing efficient catalysis of this enantiomer. In substrates of low enantioselectivity~E Ͻ 20!, the distance d~H NE Ϫ O alc ! is less than 2.0 Å, and slow-and fast-reacting enantiomers are catalyzed at similar rates. For substrates of medium enantioselectivity~20 Ͻ E Ͻ 100!, d~H NE Ϫ O alc ! is around 2.1 Å. This simple model can be applied to predict enantioselectivity of P. cepacia lipase toward a broad range of secondary alcohols.Keywords: enantioselectivity; lipase; model; molecular dynamics; Pseudomonas cepacia; secondary alcohol; stereopreference Lipase from Pseudomonas cepacia~EC 3.1.1.3! is a popular catalyst in organic synthesis~Kazlauskas & Bornscheuer, 1998! for the kinetic resolution of racemic mixtures of secondary alcohols in hydrolysis~Laumen & Schneider, 1988;Liang & Paquette, 1990;Caron & Kazlauskas, 1991;Schneider & Georgens, 1992; Bän-ziger et al., 1993;Itoh et al., 1993;Partali et al., 1993;Takano et al., 1993;Waldinger et al., 1996!, esterification~Burgess & Jennings, 1991Uejima et al., 1993;Chadha & Manohar, 1995;Gaspar & Guerrero, 1995;Hamada et al., 1996;Petschen et al., 1996!, and transesterification~Kaminska et al., 1996;Takagi et al., 1996!. The mechanistic details of the catalytic reaction of serine hydrolases~Chapus et al., 1976;! have been well investigated by quantum-chemical methods like the ab initio and density functional theory~Hu et al., 1998!, and by semiempirical methods~Monecke et al., 1998!. The reaction mechanism of ester hydrolysis~Fig. 1! starts with the formation of a Michaelis complex followed by a first transition state~19! to the first tetrahedral intermediate~2!, where the substrate is covalently linked to side-chain oxygen Og of catalytic serine. The crucial hydrogen bonds...

show abstract

Simple conformation space search protocols for the evaluation of enantioselectivity of lipases

Cited by 15 publications

References 2 publications

Rational design of enantioselective enzymes requires considerations of entropy

Rational design of enantioselective enzymes requires considerations of entropy

Contribution of both catalytic constant and Michaelis constant to CALB enantioselectivity: Use of FEP calculations for prediction studies

Stereoselectivity of Pseudomonas cepacia lipase toward secondary alcohols: A quantitative model

Contact Info

Product

Resources

About