Negatively Supercharging Cellulases Render Them Lignin-Resistant

Timothy A. Whitehead, Chandra K. Bandi, Marissa Berger, Jihyun Park, Shishir Chundawat

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

Nonspecific adsorption of cellulases to lignin hinders enzymatic biomass deconstruction. Here, we tested the hypothesis that negatively supercharging cellulases could reduce lignin inhibition. Computational design was used to negatively supercharge the surfaces of Ruminoclostridium thermocellum family 5 CelE and a CelE-family 3a carbohydrate binding module fusion. Resulting designs maintained the same expression yield, thermal stability, and nearly identical activity on soluble substrate as the wild-type proteins. Four designs showed complete lack of inhibition by lignin but with lower cellulose conversion compared to original enzymes. Increasing salt concentrations could partially rescue the activity of supercharged enzymes, supporting a mechanism of electrostatic repulsion between designs and cellulose. Results showcase a protein engineering strategy to construct highly active cellulases that are resistant to lignin-mediated inactivation, although further work is needed to understand the relationship between negative protein surface potential and activity on insoluble polysaccharides.

Original languageEnglish (US)
Pages (from-to)6247-6252
Number of pages6
JournalACS Sustainable Chemistry and Engineering
Volume5
Issue number7
DOIs
StatePublished - Jul 3 2017

Fingerprint

Cellulases
Lignin
lignin
Proteins
Cellulose
protein
cellulose
Enzymes
enzyme
Surface potential
Polysaccharides
Carbohydrates
polysaccharide
carbohydrate
Electrostatics
Biomass
Thermodynamic stability
Fusion reactions
Salts
salt

All Science Journal Classification (ASJC) codes

  • Chemistry(all)
  • Environmental Chemistry
  • Chemical Engineering(all)
  • Renewable Energy, Sustainability and the Environment

Cite this

Whitehead, Timothy A. ; Bandi, Chandra K. ; Berger, Marissa ; Park, Jihyun ; Chundawat, Shishir. / Negatively Supercharging Cellulases Render Them Lignin-Resistant. In: ACS Sustainable Chemistry and Engineering. 2017 ; Vol. 5, No. 7. pp. 6247-6252.
@article{e220522301324efe8456f6cca9634d41,
title = "Negatively Supercharging Cellulases Render Them Lignin-Resistant",
abstract = "Nonspecific adsorption of cellulases to lignin hinders enzymatic biomass deconstruction. Here, we tested the hypothesis that negatively supercharging cellulases could reduce lignin inhibition. Computational design was used to negatively supercharge the surfaces of Ruminoclostridium thermocellum family 5 CelE and a CelE-family 3a carbohydrate binding module fusion. Resulting designs maintained the same expression yield, thermal stability, and nearly identical activity on soluble substrate as the wild-type proteins. Four designs showed complete lack of inhibition by lignin but with lower cellulose conversion compared to original enzymes. Increasing salt concentrations could partially rescue the activity of supercharged enzymes, supporting a mechanism of electrostatic repulsion between designs and cellulose. Results showcase a protein engineering strategy to construct highly active cellulases that are resistant to lignin-mediated inactivation, although further work is needed to understand the relationship between negative protein surface potential and activity on insoluble polysaccharides.",
author = "Whitehead, {Timothy A.} and Bandi, {Chandra K.} and Marissa Berger and Jihyun Park and Shishir Chundawat",
year = "2017",
month = "7",
day = "3",
doi = "10.1021/acssuschemeng.7b01202",
language = "English (US)",
volume = "5",
pages = "6247--6252",
journal = "ACS Sustainable Chemistry and Engineering",
issn = "2168-0485",
publisher = "American Chemical Society",
number = "7",

}

Negatively Supercharging Cellulases Render Them Lignin-Resistant. / Whitehead, Timothy A.; Bandi, Chandra K.; Berger, Marissa; Park, Jihyun; Chundawat, Shishir.

In: ACS Sustainable Chemistry and Engineering, Vol. 5, No. 7, 03.07.2017, p. 6247-6252.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Negatively Supercharging Cellulases Render Them Lignin-Resistant

AU - Whitehead, Timothy A.

AU - Bandi, Chandra K.

AU - Berger, Marissa

AU - Park, Jihyun

AU - Chundawat, Shishir

PY - 2017/7/3

Y1 - 2017/7/3

N2 - Nonspecific adsorption of cellulases to lignin hinders enzymatic biomass deconstruction. Here, we tested the hypothesis that negatively supercharging cellulases could reduce lignin inhibition. Computational design was used to negatively supercharge the surfaces of Ruminoclostridium thermocellum family 5 CelE and a CelE-family 3a carbohydrate binding module fusion. Resulting designs maintained the same expression yield, thermal stability, and nearly identical activity on soluble substrate as the wild-type proteins. Four designs showed complete lack of inhibition by lignin but with lower cellulose conversion compared to original enzymes. Increasing salt concentrations could partially rescue the activity of supercharged enzymes, supporting a mechanism of electrostatic repulsion between designs and cellulose. Results showcase a protein engineering strategy to construct highly active cellulases that are resistant to lignin-mediated inactivation, although further work is needed to understand the relationship between negative protein surface potential and activity on insoluble polysaccharides.

AB - Nonspecific adsorption of cellulases to lignin hinders enzymatic biomass deconstruction. Here, we tested the hypothesis that negatively supercharging cellulases could reduce lignin inhibition. Computational design was used to negatively supercharge the surfaces of Ruminoclostridium thermocellum family 5 CelE and a CelE-family 3a carbohydrate binding module fusion. Resulting designs maintained the same expression yield, thermal stability, and nearly identical activity on soluble substrate as the wild-type proteins. Four designs showed complete lack of inhibition by lignin but with lower cellulose conversion compared to original enzymes. Increasing salt concentrations could partially rescue the activity of supercharged enzymes, supporting a mechanism of electrostatic repulsion between designs and cellulose. Results showcase a protein engineering strategy to construct highly active cellulases that are resistant to lignin-mediated inactivation, although further work is needed to understand the relationship between negative protein surface potential and activity on insoluble polysaccharides.

UR - http://www.scopus.com/inward/record.url?scp=85021916506&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85021916506&partnerID=8YFLogxK

U2 - 10.1021/acssuschemeng.7b01202

DO - 10.1021/acssuschemeng.7b01202

M3 - Article

AN - SCOPUS:85021916506

VL - 5

SP - 6247

EP - 6252

JO - ACS Sustainable Chemistry and Engineering

JF - ACS Sustainable Chemistry and Engineering

SN - 2168-0485

IS - 7

ER -