Synthesis of silica particles and their application as supports for alcohol dehydrogenases and cofactor immobilizations: conformational changes that lead to switch in enzyme stereoselectivity
Jazyk angličtina Země Nizozemsko Médium print-electronic
Typ dokumentu časopisecké články, práce podpořená grantem
PubMed
22472304
DOI
10.1016/j.bbapap.2012.03.010
PII: S1570-9639(12)00057-X
Knihovny.cz E-zdroje
- MeSH
- aldehydreduktasa MeSH
- aldo-keto reduktasy MeSH
- alkoholdehydrogenasa chemie MeSH
- alkoholoxidoreduktasy chemie MeSH
- cirkulární dichroismus MeSH
- enzymy imobilizované chemie MeSH
- fluorescenční spektrometrie MeSH
- játra enzymologie MeSH
- kinetika MeSH
- koenzymy chemie MeSH
- koně MeSH
- NAD chemie MeSH
- nanočástice chemie ultrastruktura MeSH
- oxid křemičitý chemie MeSH
- rozbalení proteinů MeSH
- sekundární struktura proteinů MeSH
- spektroskopie infračervená s Fourierovou transformací MeSH
- stereoizomerie MeSH
- substrátová specifita MeSH
- terciární struktura proteinů MeSH
- tryptofan chemie MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- aldehydreduktasa MeSH
- aldo-keto reduktasy MeSH
- alkoholdehydrogenasa MeSH
- alkoholoxidoreduktasy MeSH
- enzymy imobilizované MeSH
- koenzymy MeSH
- NAD MeSH
- oxid křemičitý MeSH
- tryptofan MeSH
FTIR, circular dichroism (CD) and fluorescence spectroscopies were used to characterize conformational changes in horse liver alcohol dehydrogenase (HLADH) and ketoreductase (KRED 117) upon physical and covalent immobilizations on silica particles (functionalized with amino, epoxy and thiol groups) of different sizes. Conformational changes for immobilized enzymes were associated with high and low frequency shifts of the amide I and II bands. CD spectra of native HLADH and KRED 117 characterized with a negative peak at 222nm indicating a α-helical structure. The disappearance of the negative peak in the CD spectra of immobilized enzymes and appearance of a positive peak at 222nm supported these observations. These findings demonstrated unfolding of folded enzymes and exposure of the amino acid residues during denaturation with a red shift in tryptophan fluorescence. The decrease in specific activities (by 60-70% in all cases) for both immobilized enzymes was correlated to those of conformational changes. Silica-attached enzyme-NADH systems were evaluated for enantioselective reduction of 1-(p-methoxyphenyl)-propan-2-one. Conformational changes enhanced the enantioselectivity of immobilized HLADH with a switch in its stereoselectivity. In the case of immobilized KRED 117, kinetic values (V(max) and K(m)) were lower than that of the free enzyme, without enhancing enzyme enantio- and stereoselectivity.
Biochim Biophys Acta. 2013 Aug;1834(8):1681 PubMed
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