Selective Functionalization of Microgels with Enzymes by Sortagging

Zhi Zou; Elisabeth Gau; Islam El-Awaad; Felix Jakob; Andrij Pich; Ulrich Schwaneberg

doi:10.1021/acs.bioconjchem.9b00568

Selective Functionalization of Microgels with Enzymes by Sortagging

Zhi Zou, Elisabeth Gau, Islam El-Awaad, Felix Jakob, Andrij Pich^*, Ulrich Schwaneberg^*

^*Corresponding author for this work

Research output: Contribution to journal › Article › Academic › peer-review

Abstract

Enzyme immobilization has been widely used to improve the stability and recyclability of enzymes in industrial processes. In this work, a sortase-mediated and therefore selective covalent immobilization strategy (sortagging) for enzymes on microgels (GelZyms) was investigated. Aqueous microgels were synthesized from poly(N-vinylcaprolactam)/glycidyl methacrylate (PVCL/GMA) and tagged with the sortase A recognition peptide sequence (LPETG) or its nucleophilic counterpart-tag (GGG). General applicability and selective immobilization were confirmed by subsequent sortagging of five different enzymes (Bacillus subtilis lipase A (BSLA), Yersinia mollaretii phytase (Ym-phytase), Escherichia coli copper efflux oxidase (CueO laccase), cellulase A2, and Bacillus megaterium monooxygenase P450 BM3). The latter was performed directly from the cell lysate to ensure cost-effective immobilization. All five immobilized enzymes were catalytically active and could be recycled (e.g., laccase CueO and monooxygenase P450 BM3 F87A; >55% residual activity after six cycles). Application potential was demonstrated by using CueO decorated microgels for bleaching of the synthetic dye indigo carmine.

Original language	English
Pages (from-to)	2859-2869
Number of pages	11
Journal	Bioconjugate Chemistry
Volume	30
Issue number	11
DOIs	https://doi.org/10.1021/acs.bioconjchem.9b00568
Publication status	Published - Nov 2019

Keywords

THROUGHPUT SCREENING PLATFORM
BACILLUS-SUBTILIS LIPASE
SORTASE-A
DIRECTED EVOLUTION
IMMOBILIZATION
PROTEINS
STABILITY
LACCASE
BINDING
POLY(N-VINYLCAPROLACTAM)

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10.1021/acs.bioconjchem.9b00568

Cite this

@article{5bf37f27986b401aaec1dc2d33b6ba4f,

title = "Selective Functionalization of Microgels with Enzymes by Sortagging",

abstract = "Enzyme immobilization has been widely used to improve the stability and recyclability of enzymes in industrial processes. In this work, a sortase-mediated and therefore selective covalent immobilization strategy (sortagging) for enzymes on microgels (GelZyms) was investigated. Aqueous microgels were synthesized from poly(N-vinylcaprolactam)/glycidyl methacrylate (PVCL/GMA) and tagged with the sortase A recognition peptide sequence (LPETG) or its nucleophilic counterpart-tag (GGG). General applicability and selective immobilization were confirmed by subsequent sortagging of five different enzymes (Bacillus subtilis lipase A (BSLA), Yersinia mollaretii phytase (Ym-phytase), Escherichia coli copper efflux oxidase (CueO laccase), cellulase A2, and Bacillus megaterium monooxygenase P450 BM3). The latter was performed directly from the cell lysate to ensure cost-effective immobilization. All five immobilized enzymes were catalytically active and could be recycled (e.g., laccase CueO and monooxygenase P450 BM3 F87A; >55% residual activity after six cycles). Application potential was demonstrated by using CueO decorated microgels for bleaching of the synthetic dye indigo carmine.",

keywords = "THROUGHPUT SCREENING PLATFORM, BACILLUS-SUBTILIS LIPASE, SORTASE-A, DIRECTED EVOLUTION, IMMOBILIZATION, PROTEINS, STABILITY, LACCASE, BINDING, POLY(N-VINYLCAPROLACTAM)",

author = "Zhi Zou and Elisabeth Gau and Islam El-Awaad and Felix Jakob and Andrij Pich and Ulrich Schwaneberg",

note = "Funding Information: We thank DFG Collaborative Research Center 985 “Functional Microgels and Microgel Systems” for financial support. Z.Z. gratefully acknowledges the Chinese Scholarship Council (CSC No. 201406740053) for his Ph.D. fellowship. This work was performed in parts at the Center for Chemical Polymer Technology CPT, which was supported by the EU and the federal state of North Rhine-Westphalia (Grant EFRE 300088302). Publisher Copyright: {\textcopyright} 2019 American Chemical Society.",

year = "2019",

month = nov,

doi = "10.1021/acs.bioconjchem.9b00568",

language = "English",

volume = "30",

pages = "2859--2869",

journal = "Bioconjugate Chemistry",

issn = "1043-1802",

publisher = "American Chemical Society",

number = "11",

}

TY - JOUR

T1 - Selective Functionalization of Microgels with Enzymes by Sortagging

AU - Zou, Zhi

AU - Gau, Elisabeth

AU - El-Awaad, Islam

AU - Jakob, Felix

AU - Pich, Andrij

AU - Schwaneberg, Ulrich

N1 - Funding Information: We thank DFG Collaborative Research Center 985 “Functional Microgels and Microgel Systems” for financial support. Z.Z. gratefully acknowledges the Chinese Scholarship Council (CSC No. 201406740053) for his Ph.D. fellowship. This work was performed in parts at the Center for Chemical Polymer Technology CPT, which was supported by the EU and the federal state of North Rhine-Westphalia (Grant EFRE 300088302). Publisher Copyright: © 2019 American Chemical Society.

PY - 2019/11

Y1 - 2019/11

N2 - Enzyme immobilization has been widely used to improve the stability and recyclability of enzymes in industrial processes. In this work, a sortase-mediated and therefore selective covalent immobilization strategy (sortagging) for enzymes on microgels (GelZyms) was investigated. Aqueous microgels were synthesized from poly(N-vinylcaprolactam)/glycidyl methacrylate (PVCL/GMA) and tagged with the sortase A recognition peptide sequence (LPETG) or its nucleophilic counterpart-tag (GGG). General applicability and selective immobilization were confirmed by subsequent sortagging of five different enzymes (Bacillus subtilis lipase A (BSLA), Yersinia mollaretii phytase (Ym-phytase), Escherichia coli copper efflux oxidase (CueO laccase), cellulase A2, and Bacillus megaterium monooxygenase P450 BM3). The latter was performed directly from the cell lysate to ensure cost-effective immobilization. All five immobilized enzymes were catalytically active and could be recycled (e.g., laccase CueO and monooxygenase P450 BM3 F87A; >55% residual activity after six cycles). Application potential was demonstrated by using CueO decorated microgels for bleaching of the synthetic dye indigo carmine.

AB - Enzyme immobilization has been widely used to improve the stability and recyclability of enzymes in industrial processes. In this work, a sortase-mediated and therefore selective covalent immobilization strategy (sortagging) for enzymes on microgels (GelZyms) was investigated. Aqueous microgels were synthesized from poly(N-vinylcaprolactam)/glycidyl methacrylate (PVCL/GMA) and tagged with the sortase A recognition peptide sequence (LPETG) or its nucleophilic counterpart-tag (GGG). General applicability and selective immobilization were confirmed by subsequent sortagging of five different enzymes (Bacillus subtilis lipase A (BSLA), Yersinia mollaretii phytase (Ym-phytase), Escherichia coli copper efflux oxidase (CueO laccase), cellulase A2, and Bacillus megaterium monooxygenase P450 BM3). The latter was performed directly from the cell lysate to ensure cost-effective immobilization. All five immobilized enzymes were catalytically active and could be recycled (e.g., laccase CueO and monooxygenase P450 BM3 F87A; >55% residual activity after six cycles). Application potential was demonstrated by using CueO decorated microgels for bleaching of the synthetic dye indigo carmine.

KW - THROUGHPUT SCREENING PLATFORM

KW - BACILLUS-SUBTILIS LIPASE

KW - SORTASE-A

KW - DIRECTED EVOLUTION

KW - IMMOBILIZATION

KW - PROTEINS

KW - STABILITY

KW - LACCASE

KW - BINDING

KW - POLY(N-VINYLCAPROLACTAM)

U2 - 10.1021/acs.bioconjchem.9b00568

DO - 10.1021/acs.bioconjchem.9b00568

M3 - Article

C2 - 31577418

SN - 1043-1802

VL - 30

SP - 2859

EP - 2869

JO - Bioconjugate Chemistry

JF - Bioconjugate Chemistry

IS - 11

ER -