High-resolution structural characterization of a heterogeneous biocatalyst using solid-state NMR

Sabu Varghese, Peter J. Halling, Daniel Haüssinger, Stephen Wimperis

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

Solid-state magic-angle spinning (MAS) NMR spectroscopy was employed to investigate structural detail in the enzyme human carbonic anhydrase II (hCA II) in uniformly 15N and selectively (15N leucine) enriched states, covalently immobilized on epoxy-functionalized silica. The immobilized hCA II retained 71% of its specific enzymatic activity when compared to the free enzyme in solution. On the basis of the one- and two-dimensional 1H, 13C, 15N, and 29Si MAS NMR spectra, chemical shift assignments could be obtained from the silica support, covalent linker, and immobilized enzyme. The successful covalent immobilization of the enzyme on epoxy−silica was confirmed by the appearance of signals from the aromatic and carbonyl groups in the immobilized enzyme in addition to signals from the modified support. Most notably, our MAS NMR results suggest that the covalent immobilization of the hCA II on epoxy−silica does not significantly affect the structural integrity of the protein.

LanguageEnglish
Pages28717-28726
Number of pages10
JournalJournal of Physical Chemistry C
Volume120
Issue number50
Early online date25 Nov 2016
DOIs
Publication statusPublished - 22 Dec 2016

Fingerprint

Biocatalysts
Carbonic Anhydrase II
Silicon Dioxide
Magic angle spinning
Carbonic anhydrase
enzymes
carbonic anhydrase
Enzymes
Nuclear magnetic resonance
solid state
Immobilized Enzymes
nuclear magnetic resonance
Silica
high resolution
metal spinning
silicon dioxide
immobilization
Catalyst supports
Chemical shift
Structural integrity

Keywords

  • solid state NMR spectroscopy
  • protein
  • immobilized enzymes

Cite this

Varghese, Sabu ; Halling, Peter J. ; Haüssinger, Daniel ; Wimperis, Stephen. / High-resolution structural characterization of a heterogeneous biocatalyst using solid-state NMR. In: Journal of Physical Chemistry C. 2016 ; Vol. 120, No. 50. pp. 28717-28726.
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High-resolution structural characterization of a heterogeneous biocatalyst using solid-state NMR. / Varghese, Sabu; Halling, Peter J.; Haüssinger, Daniel; Wimperis, Stephen.

In: Journal of Physical Chemistry C, Vol. 120, No. 50, 22.12.2016, p. 28717-28726.

Research output: Contribution to journalArticle

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