Detecting Cellulase Penetration Into Corn Stover Cell Walls by Immuno-Electron Microscopy
Abstract
In general, pretreatments are designed to enhance the accessibility of cellulose to enzymes, allowing for more efficient conversion. In this study, we have detected the penetration of major cellulases present in a commercial enzyme preparation (Spezyme CP) into corn stem cell walls following mild-, moderate- and high-severity dilute sulfuric acid pretreatments. The Trichoderma reesei enzymes, Cel7A (CBH I) and Cel7B (EG I), as well as the cell wall matrix components xylan and lignin were visualized within digested corn stover cell walls by immuno transmission electron microscopy (TEM) using enzyme- and polymer-specific antibodies. Low severity dilute-acid pretreatment (20 min at 100 C) enabled <1% of the thickness of secondary cell walls to be penetrated by enzyme, moderate severity pretreatment at (20 min at 120 C) allowed the enzymes to penetrate {approx}20% of the cell wall, and the high severity (20 min pretreatment at 150 C) allowed 100% penetration of even the thickest cell walls. These data allow direct visualization of the dramatic effect dilute-acid pretreatment has on altering the condensed ultrastructure of biomass cell walls. Loosening of plant cell wall structure due to pretreatment and the subsequently improved access by cellulases has been hypothesized by the biomass conversion community for overmore »
- Authors:
- Publication Date:
- Research Org.:
- National Renewable Energy Lab. (NREL), Golden, CO (United States)
- Sponsoring Org.:
- USDOE
- OSTI Identifier:
- 969729
- DOE Contract Number:
- AC36-99-GO10337
- Resource Type:
- Journal Article
- Journal Name:
- Biotechnology and Bioengineering
- Additional Journal Information:
- Journal Volume: 103; Journal Issue: 3, 15 June 2009; Journal ID: ISSN 0006-3592
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 09 BIOMASS FUELS; 59 BASIC BIOLOGICAL SCIENCES; AGRICULTURAL WASTES; ANTIBODIES; BIOMASS; CELL WALL; CELLULASE; CELLULOSE; ENZYMES; HYPOTHESIS; LIGNIN; MAIZE; MICROSCOPY; PLANT CELLS; STEM CELLS; SULFURIC ACID; THICKNESS; TRANSMISSION ELECTRON MICROSCOPY; TRICHODERMA VIRIDE; XYLANS; Chemical and Biosciences
Citation Formats
Donohoe, B S, Selig, M J, Viamajala, S, Vinzant, T B, Adney, W S, and Himmel, M E. Detecting Cellulase Penetration Into Corn Stover Cell Walls by Immuno-Electron Microscopy. United States: N. p., 2009.
Web. doi:10.1002/bit.22281.
Donohoe, B S, Selig, M J, Viamajala, S, Vinzant, T B, Adney, W S, & Himmel, M E. Detecting Cellulase Penetration Into Corn Stover Cell Walls by Immuno-Electron Microscopy. United States. https://doi.org/10.1002/bit.22281
Donohoe, B S, Selig, M J, Viamajala, S, Vinzant, T B, Adney, W S, and Himmel, M E. 2009.
"Detecting Cellulase Penetration Into Corn Stover Cell Walls by Immuno-Electron Microscopy". United States. https://doi.org/10.1002/bit.22281.
@article{osti_969729,
title = {Detecting Cellulase Penetration Into Corn Stover Cell Walls by Immuno-Electron Microscopy},
author = {Donohoe, B S and Selig, M J and Viamajala, S and Vinzant, T B and Adney, W S and Himmel, M E},
abstractNote = {In general, pretreatments are designed to enhance the accessibility of cellulose to enzymes, allowing for more efficient conversion. In this study, we have detected the penetration of major cellulases present in a commercial enzyme preparation (Spezyme CP) into corn stem cell walls following mild-, moderate- and high-severity dilute sulfuric acid pretreatments. The Trichoderma reesei enzymes, Cel7A (CBH I) and Cel7B (EG I), as well as the cell wall matrix components xylan and lignin were visualized within digested corn stover cell walls by immuno transmission electron microscopy (TEM) using enzyme- and polymer-specific antibodies. Low severity dilute-acid pretreatment (20 min at 100 C) enabled <1% of the thickness of secondary cell walls to be penetrated by enzyme, moderate severity pretreatment at (20 min at 120 C) allowed the enzymes to penetrate {approx}20% of the cell wall, and the high severity (20 min pretreatment at 150 C) allowed 100% penetration of even the thickest cell walls. These data allow direct visualization of the dramatic effect dilute-acid pretreatment has on altering the condensed ultrastructure of biomass cell walls. Loosening of plant cell wall structure due to pretreatment and the subsequently improved access by cellulases has been hypothesized by the biomass conversion community for over two decades, and for the first time, this study provides direct visual evidence to verify this hypothesis. Further, the high-resolution enzyme penetration studies presented here provide insight into the mechanisms of cell wall deconstruction by cellulolytic enzymes.},
doi = {10.1002/bit.22281},
url = {https://www.osti.gov/biblio/969729},
journal = {Biotechnology and Bioengineering},
issn = {0006-3592},
number = 3, 15 June 2009,
volume = 103,
place = {United States},
year = {Mon Jun 15 00:00:00 EDT 2009},
month = {Mon Jun 15 00:00:00 EDT 2009}
}