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Title: Impacts of cellulase deactivation at the moving air–liquid interface on cellulose conversions at low enzyme loadings

Abstract

Background: We recently confirmed that the deactivation of T. reesei cellulases at the air–liquid interface reduces microcrystalline cellulose conversion at low enzyme loadings in shaken flasks. It is one of the main causes for lowering of cellulose conversions at low enzyme loadings. However, supplementing cellulases with small quantities of surface-active additives in shaken flasks can increase cellulose conversions at low enzyme loadings. It was also shown that cellulose conversions at low enzyme loadings can be increased in unshaken flasks if the reactions are carried for a longer time. This study further explores these recent findings to better understand the impact of air–liquid interfacial phenomena on enzymatic hydrolysis of cellulose contained in Avicel, Sigmacell, α-cellulose, cotton linters, and filter paper. The impacts of solids and enzyme loadings, supplementation with nonionic surfactant Tween 20 and xylanases, and application of different types of mixing and reactor designs on cellulose hydrolysis were also evaluated.Results: Avicel cellulose conversions at high solid loading were more than doubled by minimizing loss of cellulases to the air–liquid interface. Maximum cellulose conversions were high for surface-active supplemented shaken flasks or unshaken flasks because of low cellulase deactivation at the air–liquid interface. The nonionic surfactant Tween 20 was unable tomore » completely prevent cellulase deactivation in shaken flasks and only reduced cellulose conversions at unreasonably high concentrations.Conclusions: High dynamic interfacial areas created through baffles in reactor vessels, low volumes in high-capacity vessels, or high shaking speeds severely limited cellulose conversions at low enzyme loadings. Precipitation of cellulases due to aggregation at the air–liquid interface caused their continuous deactivation in shaken flasks and severely limited solubilization of cellulose.« less

Authors:
 [1]; ORCiD logo [2];  [3]
  1. Univ. of California, Riverside, CA (United States). Bourns College of Engineering, Center for Environmental Research and Technology (CE‑CERT) and Center for Environmental Research and Technology (CE‑CERT); Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). BioEnergy Science Center (BESC)
  2. Univ. of California, Riverside, CA (United States). Bourns College of Engineering, Center for Environmental Research and Technology (CE‑CERT) and Center for Environmental Research and Technology (CE‑CERT); Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). BioEnergy Science Center (BESC) and Center for Bioenergy Innovation (CBI)
  3. Univ. of California, Riverside, CA (United States). Bourns College of Engineering, Center for Environmental Research and Technology (CE‑CERT); Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). BioEnergy Science Center (BESC) and Center for Bioenergy Innovation (CBI)
Publication Date:
Research Org.:
Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
Sponsoring Org.:
USDOE
OSTI Identifier:
1511911
Grant/Contract Number:  
AC05-00OR22725
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
Biotechnology for Biofuels
Additional Journal Information:
Journal Volume: 12; Journal Issue: 1; Journal ID: ISSN 1754-6834
Publisher:
BioMed Central
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; Cellulose; Cellulase; Deactivation; Hydrolysis; Air–liquid interface; Gas–liquid interface

Citation Formats

Bhagia, Samarthya, Wyman, Charles E., and Kumar, Rajeev. Impacts of cellulase deactivation at the moving air–liquid interface on cellulose conversions at low enzyme loadings. United States: N. p., 2019. Web. doi:10.1186/s13068-019-1439-2.
Bhagia, Samarthya, Wyman, Charles E., & Kumar, Rajeev. Impacts of cellulase deactivation at the moving air–liquid interface on cellulose conversions at low enzyme loadings. United States. doi:10.1186/s13068-019-1439-2.
Bhagia, Samarthya, Wyman, Charles E., and Kumar, Rajeev. Tue . "Impacts of cellulase deactivation at the moving air–liquid interface on cellulose conversions at low enzyme loadings". United States. doi:10.1186/s13068-019-1439-2. https://www.osti.gov/servlets/purl/1511911.
@article{osti_1511911,
title = {Impacts of cellulase deactivation at the moving air–liquid interface on cellulose conversions at low enzyme loadings},
author = {Bhagia, Samarthya and Wyman, Charles E. and Kumar, Rajeev},
abstractNote = {Background: We recently confirmed that the deactivation of T. reesei cellulases at the air–liquid interface reduces microcrystalline cellulose conversion at low enzyme loadings in shaken flasks. It is one of the main causes for lowering of cellulose conversions at low enzyme loadings. However, supplementing cellulases with small quantities of surface-active additives in shaken flasks can increase cellulose conversions at low enzyme loadings. It was also shown that cellulose conversions at low enzyme loadings can be increased in unshaken flasks if the reactions are carried for a longer time. This study further explores these recent findings to better understand the impact of air–liquid interfacial phenomena on enzymatic hydrolysis of cellulose contained in Avicel, Sigmacell, α-cellulose, cotton linters, and filter paper. The impacts of solids and enzyme loadings, supplementation with nonionic surfactant Tween 20 and xylanases, and application of different types of mixing and reactor designs on cellulose hydrolysis were also evaluated.Results: Avicel cellulose conversions at high solid loading were more than doubled by minimizing loss of cellulases to the air–liquid interface. Maximum cellulose conversions were high for surface-active supplemented shaken flasks or unshaken flasks because of low cellulase deactivation at the air–liquid interface. The nonionic surfactant Tween 20 was unable to completely prevent cellulase deactivation in shaken flasks and only reduced cellulose conversions at unreasonably high concentrations.Conclusions: High dynamic interfacial areas created through baffles in reactor vessels, low volumes in high-capacity vessels, or high shaking speeds severely limited cellulose conversions at low enzyme loadings. Precipitation of cellulases due to aggregation at the air–liquid interface caused their continuous deactivation in shaken flasks and severely limited solubilization of cellulose.},
doi = {10.1186/s13068-019-1439-2},
journal = {Biotechnology for Biofuels},
issn = {1754-6834},
number = 1,
volume = 12,
place = {United States},
year = {2019},
month = {4}
}

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    Works referencing / citing this record:

    Effects of the surfactant tween 80 on enzymatic hydrolysis of newspaper
    journal, June 1981

    • Castanon, Marisi; Wilke, Charle R.
    • Biotechnology and Bioengineering, Vol. 23, Issue 6
    • DOI: 10.1002/bit.260230615

    Effects of shear on proteins in solution
    journal, November 2010


    Inhibition ofTrichoderma reesei cellulase by sugars and solvents
    journal, July 1990

    • Holtzapple, Mark; Cognata, Mona; Shu, Yuancai
    • Biotechnology and Bioengineering, Vol. 36, Issue 3
    • DOI: 10.1002/bit.260360310

    Why proteins prefer interfaces
    journal, January 1991


    Non-ionic surfactants do not consistently improve the enzymatic hydrolysis of pure cellulose
    journal, April 2015


    Techno-economic comparison of process technologies for biochemical ethanol production from corn stover
    journal, November 2010


    Surface tension of bovine serum albumin and tween 20 at the air-aqueous interface
    journal, October 1998

    • NiɁo, Ma Rosario Rodríguez; Patino, J. M. Rodríguez
    • Journal of the American Oil Chemists' Society, Vol. 75, Issue 10
    • DOI: 10.1007/s11746-998-0169-6

    A simple method for displaying the hydropathic character of a protein
    journal, May 1982


    Yield-determining factors in high-solids enzymatic hydrolysis of lignocellulose
    journal, January 2009

    • Kristensen, Jan B.; Felby, Claus; Jørgensen, Henning
    • Biotechnology for Biofuels, Vol. 2, Issue 1
    • DOI: 10.1186/1754-6834-2-11

    Effect of endoxylanase and α-l-arabinofuranosidase supplementation on the enzymatic hydrolysis of steam exploded wheat straw
    journal, March 2011


    Inhibition and stimulation of the cellulase of Myrothecium verrucaria
    journal, January 1953


    Do Protein Molecules Unfold in a Simple Shear Flow?
    journal, November 2006


    Biochemical characterization and synergism of cellulolytic enzyme system from Chaetomium globosum on rice straw saccharification
    journal, November 2016

    • Wanmolee, Wanwitoo; Sornlake, Warasirin; Rattanaphan, Nakul
    • BMC Biotechnology, Vol. 16, Issue 1
    • DOI: 10.1186/s12896-016-0312-7

    An Unfavourable Effect of Shaking on Fungal Cellulases
    journal, August 1956


    An Effect of Proteins and Proteoses on the Cellulase of Myrothecium verrucaria
    journal, July 1952


    Proteins at liquid interfaces
    journal, July 1979


    Surface deactivation of cellulase and its prevention
    journal, March 1982


    Deactivation of Cellulase at the Air-Liquid Interface Is the Main Cause of Incomplete Cellulose Conversion at Low Enzyme Loadings
    journal, January 2018


    What is (and is not) vital to advancing cellulosic ethanol
    journal, April 2007


    Structure of Cotton Linters
    journal, August 1947


    Measurement of protein using bicinchoninic acid
    journal, October 1985


    Strong cellulase inhibition by Mannan polysaccharides in cellulose conversion to sugars: Strong Cellulase Inhibition by Heteromannans
    journal, May 2014

    • Kumar, Rajeev; Wyman, Charles E.
    • Biotechnology and Bioengineering, Vol. 111, Issue 7
    • DOI: 10.1002/bit.25218

    Orogenic Displacement of Protein from the Air/Water Interface by Competitive Adsorption
    journal, February 1999

    • Mackie, Alan R.; Gunning, A. Patrick; Wilde, Peter J.
    • Journal of Colloid and Interface Science, Vol. 210, Issue 1
    • DOI: 10.1006/jcis.1998.5941

    Evaluation of several microcrystalline celluloses obtained from agricultural by-products
    journal, January 2011

    • Rojas, John; Lopez, Alvin; Guisao, Santiago
    • Journal of Advanced Pharmaceutical Technology & Research, Vol. 2, Issue 3
    • DOI: 10.4103/2231-4040.85527

    Kinetics of adsorption of globular proteins at an air-water interface
    journal, May 1992

    • Narsimhan, Ganesan; Uraizee, Farooq
    • Biotechnology Progress, Vol. 8, Issue 3
    • DOI: 10.1021/bp00015a003

    Design of reaction systems for specialty organic chemicals
    journal, January 1988


    Kinetics of protein unfolding at interfaces
    journal, November 2012


    Robustness of two-step acid hydrolysis procedure for composition analysis of poplar
    journal, September 2016


    Estimation of diffusion coefficients of proteins
    journal, May 1980

    • Young, M. E.; Carroad, P. A.; Bell, R. L.
    • Biotechnology and Bioengineering, Vol. 22, Issue 5
    • DOI: 10.1002/bit.260220504

    Harnessing the potential of LPMO-containing cellulase cocktails poses new demands on processing conditions
    journal, November 2015

    • Müller, Gerdt; Várnai, Anikó; Johansen, Katja Salomon
    • Biotechnology for Biofuels, Vol. 8, Issue 1
    • DOI: 10.1186/s13068-015-0376-y

    Access of cellulase to cellulose and lignin for poplar solids produced by leading pretreatment technologies
    journal, May 2009

    • Kumar, Rajeev; Wyman, Charles E.
    • Biotechnology Progress, Vol. 25, Issue 3
    • DOI: 10.1002/btpr.153

    Protein–surfactant interactions: A tale of many states
    journal, May 2011


    Cellulase adsorption and relationship to features of corn stover solids produced by leading pretreatments
    journal, June 2009

    • Kumar, Rajeev; Wyman, Charles E.
    • Biotechnology and Bioengineering, Vol. 103, Issue 2
    • DOI: 10.1002/bit.22258

    Substrate and Enzyme Characteristics that Limit Cellulose Hydrolysis
    journal, October 1999

    • Mansfield, S. D.; Mooney, C.; Saddler, J. N.
    • Biotechnology Progress, Vol. 15, Issue 5
    • DOI: 10.1021/bp9900864

    A mechanistic model of the enzymatic hydrolysis of cellulose
    journal, May 2010

    • Levine, Seth E.; Fox, Jerome M.; Blanch, Harvey W.
    • Biotechnology and Bioengineering, Vol. 107, Issue 1
    • DOI: 10.1002/bit.22789

    Kinetic analysis of bioconversion of cellulose in attrition bioreactor
    journal, January 1988

    • Jones, Evan O.; Lee, James M.
    • Biotechnology and Bioengineering, Vol. 31, Issue 1
    • DOI: 10.1002/bit.260310106

    Shear Deactivation of Cellulase, Exoglucanase, Endoglucanase, and β-Glucosidase in a Mechanically Agitated Reactor
    journal, December 2001

    • Gunjikar, T. P.; Sawant, S. B.; Joshi, J. B.
    • Biotechnology Progress, Vol. 17, Issue 6
    • DOI: 10.1021/bp010114u

    Measurement of cellulase activities
    journal, January 1987


    An improved method to directly estimate cellulase adsorption on biomass solids
    journal, April 2008


    Enhancement of enzymatic hydrolysis of cellulose by surfactant
    journal, November 1986

    • Ooshima, Hiroshi; Sakata, Masaru; Harano, Yoshio
    • Biotechnology and Bioengineering, Vol. 28, Issue 11
    • DOI: 10.1002/bit.260281117

    The effect of bovine serum albumin on batch and continuous enzymatic cellulose hydrolysis mixed by stirring or shaking
    journal, May 2011