Creating Space for Large Secondary Alcohols by Rational Redesign of <i>Candida antarctica</i> Lipase B

Magnusson, Anders O.; Rotticci-Mulder, Johanna C.; Santagostino, Alberto; Hult, Karl

doi:10.1002/cbic.200400410

Cited by 108 publications

(64 citation statements)

References 16 publications

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“…[26][27][28] This enzyme was readily bound to EziG™ within 30 min of incubation while ten days of incubation was required for immobilization on Accurel®. For this variant, EziG™ (HybCPG VBC) is a more suitable carrier than Accurel® based on the bulk volumetric activity and the time needed for immobilization (see Table S2, Supplementary Information).…”

Section: Methodsmentioning

confidence: 99%

A general protein purification and immobilization method on controlled porosity glass: biocatalytic applications

et al. 2014

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Section: Methodsmentioning

confidence: 99%

A general protein purification and immobilization method on controlled porosity glass: biocatalytic applications

et al. 2014

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“…[20] Similarly, a W104A mutant expanded this pocket and showed increased activity with alcohols such as heptan-4-ol and nonan-5-ol. [21] A C H T U N G T R E N N U N G Interestingly, Hult and co-workers have also investigated aldol and Michael additions, [22][23][24][25] and Baeyer-Villiger oxidations [26] with the catalytic serine variant S105A. In a novel approach,…”

Section: Introductionmentioning

confidence: 97%

Understanding the Plasticity of the α/β Hydrolase Fold: Lid Swapping on the Candida antarctica Lipase B Results in Chimeras with Interesting Biocatalytic Properties

et al. 2009

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The best of both worlds. Long molecular dynamics (MD) simulations of Candida antarctica lipase B (CALB) confirmed the function of helix alpha5 as a lid structure. Replacement of the helix with corresponding lid regions from CALB homologues from Neurospora crassa and Gibberella zeae resulted in new CALB chimeras with novel biocatalytic properties. The figure shows a snapshot from the MD simulation. The Candida antarctica lipase B (CALB) has found very extensive use in biocatalysis reactions. Long molecular dynamics simulations of CALB in explicit aqueous solvent confirmed the high mobility of the regions lining the channel that leads into the active site, in particular, of helices alpha5 and alpha10. The simulation also confirmed the function of helix alpha5 as a lid of the lipase. Replacing it with corresponding lid regions from the CALB homologues from Neurospora crassa and Gibberella zeae resulted in two new CALB mutants. Characterization of these revealed several interesting properties, including increased hydrolytic activity on simple esters, specifically substrates with C(alpha) branching on the carboxylic side, and much increased enantioselectivity in hydrolysis of racemic ethyl 2-phenylpropanoate (E>50), which is a common structure of the profen drug family.

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“…As a matter of fact, even reversal of substrate specificity (8,9) and enantiopreference (10,11) has been reported. A higher thermostability and altered stereoselectivity for CALB in organic solvents have also been observed (12,13). Many researchers have put effort into elucidating the underlying mechanisms responsible for the observed solvent effects.…”

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Analysis of the Conformational Stability and Activity of Candida antarctica Lipase B in Organic Solvents

Cong

Tan

Zhang

et al. 2010

Journal of Biological Chemistry

149

117

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The conformational stability and activity of Candida antarctica lipase B (CALB) in the polar and nonpolar organic solvents were investigated by molecular dynamics and quantum mechanics/molecular mechanics simulations. The conformation change of CALB in the polar and nonpolar solvents was examined in two aspects: the overall conformation change of CALB and the conformation change of the active site. The simulation results show that the overall conformation of CALB is stable in the organic solvents. In the nonpolar solvents, the conformation of the active site keeps stable, whereas in the polar solvents, the solvent molecules reach into the active site and interact intensively with the active site. This interaction destroys the hydrogen bonding between Ser 105 and His 224 . In the solvents, the activation energy of CALB and that of the active site region were further simulated by quantum mechanics/molecular mechanics simulation. The results indicate that the conformation change in the region of active sites is the main factor that influences the activity of CALB.Because of its high enantioselectivity and catalytic activity, wide range of substrates, and thermal stability, Candida antarctica lipase B (CALB) 3 is widely used in many industrial applications and scientific researches (1). CALB is composed of 317 amino acid residues and has a molecular mass of 33 K (2). Similar to other serine hydrolyzes, a serine-histidine-asparate catalytic triad is responsible for the catalytic activity of CALB. The mechanism is outlined in Fig. 1. It is a two-step mechanism with an acylation step and a deacylation step separated by a covalent acyl-enzyme intermediate (3).The activity of the enzymes is strongly affected by the choice of solvent (4 -7). As a matter of fact, even reversal of substrate specificity (8, 9) and enantiopreference (10, 11) has been reported. A higher thermostability and altered stereoselectivity for CALB in organic solvents have also been observed (12, 13). Many researchers have put effort into elucidating the underlying mechanisms responsible for the observed solvent effects. The most widely accepted model was described by Laane (14), who summarized the influence of organic solvents on the enzymatic reactions and concluded that the enzyme activity is higher in the environment surrounded by nonpolar and midpolar solvents, whereas the lowest activity is expressed in polar solvents. The Laane model has been widely used in solvent selection in enzymatic reactions. However, the Laane model does not describe the mechanisms of solvent effect on a molecular level.Molecular dynamics simulations have been proven to be a useful tool in understanding protein structure and have been used to get insights into the structure and behavior of the enzymes (15-18). The effect of solvents on the activity of CALB might be the results of the conformational change around the activity site or some particular area. In this work, the overall conformational change of CALB and the local conformational change around the active site in ...

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Creating Space for Large Secondary Alcohols by Rational Redesign of Candida antarctica Lipase B

Cited by 108 publications

References 16 publications

A general protein purification and immobilization method on controlled porosity glass: biocatalytic applications

A general protein purification and immobilization method on controlled porosity glass: biocatalytic applications

Understanding the Plasticity of the α/β Hydrolase Fold: Lid Swapping on the Candida antarctica Lipase B Results in Chimeras with Interesting Biocatalytic Properties

Analysis of the Conformational Stability and Activity of Candida antarctica Lipase B in Organic Solvents

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