| 1. |
- Holmquist, Mats Torsten, et al.
(författare)
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Lipases from Rhizomucor miehei and Humicola lanuginosa : Modification of the lid covering the active site alters enantioselectivity
- 1993
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Ingår i: Journal of Protein Chemistry. - 0277-8033. ; 12:6, s. 749-757
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Tidskriftsartikel (refereegranskat)abstract
- The homologous lipases from Rhizomucor miehei and Humicola lanuginosa showed approximately the same enantioselectivity when 2-methyldecanoic acid esters were used as substrates. Both lipases preferentially hydrolyzed the S- enantiomer of 1-heptyl 2-methyldecanoate (R. miehei: E(S) = 8.5; H. lanuginosa: E(S) = 10.5), but the R-enantiomer of phenyl 2-methyldecanoate (E(R) = 2.9). Chemical arginine specific modification of the R. miehei lipase with 1,2-cyclohexanedione resulted in a decreased enantioselectivity (E(R) = 2.0), only when the phenyl ester was used as a substrate. In contrast, treatment with phenylglyoxal showed a decreased enantioselectivity (E(S) = 2.5) only when the heptyl ester was used as a substrate. The presence of guanidine, an arginine side chain analog, decreased the enantioselectivity with the heptyl ester (E(S) = 1.9) and increased the enantioselectivity with the aromatic ester (E(R) = 4.4) as substrates. The mutation, Glu 87 Ala, in the lid of the H. lanuginosa lipase, which might decrease the electrostatic stabilization of the open-lid conformation of the lipase, resulted in 47% activity compared to the native lipase, in a tributyrin assay. The Glu 87 Ala mutant showed an increased enantioselectivity with the heptyl ester (E(S) = 17.4) and a decreased enantioselectivity with the phenyl ester (E(R) = 2.5) as substrates, compared to native lipase. The enantioselectivities of both lipases in the esterification of 2-methyldecanoic acid with 1-heptanol were unaffected by the lid modifications.
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| 2. |
- Berglund, Per, et al.
(författare)
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2-Methylalkanoic acids resolved by esterification catalysed by lipase from Candida rugosa : Alcohol chain length and enantioselectivity
- 1993
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Ingår i: Tetrahedron Asymmetry. - 0957-4166. ; 4:8, s. 1869-1878
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Tidskriftsartikel (refereegranskat)abstract
- Enantiomerically pure (R)-2-methyldecanoic acid and (S)-2-methyl-1-decanol were prepared in a multi gram scale by esterification reactions catalysed by lipase from Candida rugosa. The enantiomeric ratios (E-values) were determined as a function of the chain length of the alcohol used as the complementary substrate in cyclohexane. In the resolution of 2-methyldecanoic acid the highest value (E = 37 ± 5) was obtained, when either 2-hexanol, 1-heptanol or 1-octanol were used. In contrast, when resolving 2-methyloctanoic acid, the E-values increased continually with increasing chain length of the alcohol used. 1-Hexadecanol gave the highest value: E > 100. The E-values were determined from the enantiomeric excess (ee) of the product at a conversion below 0.4. After two consecutive esterification reactions enantiomerically pure (R)-2-methyldecanoic acid, >99.8% ee, and after subsequent reduction of the ester produced, (S)-2-methyl-1-decanol, 96.7% ee, were obtained.
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| 3. |
- Berglund, Per, et al.
(författare)
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Reversed enantiopreference of Candida rugosa lipase supports different modes of binding enantiomers of a chiral acyl donor
- 1998
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Ingår i: Journal of Molecular Catalysis - B Enzymatic. - 1381-1177 .- 1873-3158. ; 5:1-4, s. 283-287
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Tidskriftsartikel (refereegranskat)abstract
- Molecular modelling identifies two different productive modes of binding the enantiomers of a 2-methyldecanoic acid ester to the active site of Candida rugosa lipase (CRL). The fast reacting S-enantiomer occupies the previously identified acyl-binding tunnel of the enzyme, whereas the R- enantiomer leaves the tunnel empty. The modelling suggested that if both enantiomers were forced to bind to the active site leaving the tunnel empty, the enzyme would reverse its enantiopreference to become R-enantioselective. To test this hypothesis, we designed a structural analogue to 2- methyldecanoic acid, 2-methyl-6-(2-thienyl)hexanoic acid, which was expected to be too bulky to fit its acyl moiety into the acyl-binding tunnel. The CRL- catalysed hydrolysis of the ethyl ester of this substrate resulted in the preferential conversion of the R-enantiomer as predicted by molecular modelling. This represents the first kinetic evidence supporting the existence of two different modes of binding the enantiomers of a 2- methyldecanoic acid ester to the active site of CRL. We have shown that a rational 3D based approach in combination with substrate engineering can be used to predict and control the stereochemical outcome of a lipase catalysed reaction.
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| 4. |
- Holmberg, Erland, et al.
(författare)
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Reaction conditions for the resolution of 2-methylalkanoic acids in esterification and hydrolysis with lipase from Candida cylindracea
- 1991
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Ingår i: Applied Microbiology and Biotechnology. - 0175-7598. ; 35:5, s. 572-578
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Tidskriftsartikel (refereegranskat)abstract
- We have demonstrated resolution of 2-methylalkanoic acids using lipase from Candida cylindracea as a catalyst. The resolution of 2-methyldecanoic acid was more successful than that of 2-methylbutyric acid both by esterification and hydrolysis. This indicates that the resolution of the acid is dependent on the chain length of the acid moiety. The chain length of the alcohol moiety of the ester affected the resolution of the long-chain acid only. Using esterification, (R)-2-methyldecanoic acid was produced in an enantiomeric excess (e.e.) of 95% (E = 40). If the enantiomeric ratio is low (E = 3.6), as in the resolution of 2-methylbutyric acid, esterification combined with a high equilibrium conversion could be used to yield the remaining acid in a high e.e. In the hydrolytic reactions, the e.e. and the equilibrium conversion were dependent on the pH and the presence of CaCl2. When octyl 2-methyldecanoate was hydrolysed at pH 8.0 in the presence of CaCl2, the (S)-acid was formed with an e.e. of 80% (E = 9), but when the hydrolysis was carried out at pH 7.5 without CaCl2, a very low e.e. and a low equilibrium conversion were observed. The latter conditions allowed the esterification of 2-methyldecanoic acid with 1-octanol even in aqueous medium.
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