2024-03-29T10:51:24Zhttp://digital.csic.es/dspace-oai/requestoai:digital.csic.es:10261/643902016-02-17T11:12:29Zcom_10261_79com_10261_1col_10261_332
DIGITAL.CSIC
author
Sandoval, Manuel
author
Cortés Cabrera, Álvaro
author
Civera, Concepción
author
Treviño, Juan
author
Ferreras, Eloy
author
Vaultier, Michel
author
Berenguer, José
author
Lozano, Pedro
author
Hernáiz, María J.
funder
Ministerio de Ciencia e Innovación (España)
funder
Universidad Complutense de Madrid
funder
Universidad de Costa Rica
2013-01-18T08:49:13Z
2013-01-18T08:49:13Z
2012
RSC Advances 2:6306-6314 (2012)
2046-2069
http://hdl.handle.net/10261/64390
10.1039/C2RA20618H
http://dx.doi.org/10.13039/501100004837http://dx.doi.org/10.13039/501100002911http://dx.doi.org/10.13039/501100005298
Room temperature ionic liquids (ILs) can affect enzyme activity in some enzyme-catalyzed reactions, however the effects of these cosolvents on the enzymes are not clearly understood. Using β-galactosidase from Thermus thermophilus HB27 (TTP0042), we found an important change from the classical regioselectivity of the transglycosylation reaction with this enzyme. The enzyme increases N-acetyl-D-lactosamine synthesis (Galβ[1→4]GlcNAc) when RTILs are used instead of the traditional self-condensated products. To understand the possible effect of these liquids on the synthetic behavior of the enzyme, we performed a molecular interaction study by surface plasmon resonance. The KD value obtained for this interaction could mean that ILs bind to β-galactosidase through non specific interactions characterized by very fast kinetics and millimolar affinity. Then, several reactions were performed, increasing the concentration of the IL. As a result, a dependence on the IL's concentration was found for transglycosylation products. We hypothesize that ILs might induce conformational changes in the enzyme, which would modify the enzymatic activity and regioselectivity. These structural modifications were confirmed in the secondary and tertiary structures of the protein by circular dichroism and fluorescence studies, respectively. Molecular modeling confirms this hypothesis and shows that the enzyme becomes more flexible in an IL–water mixture and that it allows stabilization of the GlcNAc molecule in the active centre of the enzyme, in order to develop a new product according to the original regioselectivity of the reaction.
eng
closedAccess
N-acetyl-lactosamine
Ionic liquid
Enzymatic synthesis
Thermus Thermophilus
Efficient and selective enzymatic synthesis of N-acetyl-lactosamine in ionic liquid: a rational explanation
artículo
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
URL
https://digital.csic.es/bitstream/10261/64390/1/accesoRestringido.pdf
File
MD5
42637ae8545636bc41605c1740a9a84e
15753
application/pdf
accesoRestringido.pdf
URL
https://digital.csic.es/bitstream/10261/64390/4/accesoRestringido.pdf.txt
File
MD5
7c821e3180e847b607957fd9795236ad
401
text/plain
accesoRestringido.pdf.txt