Comparative community proteomics demonstrates the unexpected importance of actinobacterial glycoside hydrolase family 12 protein for crystalline cellulose hydrolysis

Jennifer Hiras, Yu Wei Wu, Kai Deng, Carrie D. Nicora, Joshua T. Aldrich, Dario Frey, Sebastian Kolinko, Errol W. Robinson, Jon M. Jacobs, Paul D. Adams, Trent R. Northen, Blake A. Simmons, Steven W. Singer

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

Glycoside hydrolases (GHs) are key enzymes in the depolymerization of plant-derived cellulose, a process central to the global carbon cycle and the conversion of plant biomass to fuels and chemicals. A limited number of GH families hydrolyze crystalline cellulose, often by a processive mechanism along the cellulose chain. During cultivation of thermophilic cellulolytic microbial communities, substantial differences were observed in the crystalline cellulose saccharification activities of supernatants recovered from divergent lineages. Comparative community proteomics identified a set of cellulases from a population closely related to actinobacterium Thermobispora bispora that were highly abundant in the most active consortium. Among the cellulases from T. bispora, the abundance of a GH family 12 (GH12) protein correlated most closely with the changes in crystalline cellulose hydrolysis activity. This result was surprising since GH12 proteins have been predominantly characterized as enzymes active on soluble polysaccharide substrates. Heterologous expression and biochemical characterization of the suite of T. bispora hydrolytic cellulases confirmed that the GH12 protein possessed the highest activity on multiple crystalline cellulose substrates and demonstrated that it hydrolyzes cellulose chains by a predominantly random mechanism. This work suggests that the role of GH12 proteins in crystalline cellulose hydrolysis by cellulolytic microbes should be reconsidered. IMPORTANCE Cellulose is the most abundant organic polymer on earth, and its enzymatic hydrolysis is a key reaction in the global carbon cycle and the conversion of plant biomass to biofuels. The glycoside hydrolases that depolymerize crystalline cellulose have been primarily characterized from isolates. In this study, we demonstrate that adapting microbial consortia from compost to grow on crystalline cellulose generated communities whose soluble enzymes exhibit differential abilities to hydrolyze crystalline cellulose. Comparative proteomics of these communities identified a protein of glycoside hydrolase family 12 (GH12), a family of proteins previously observed to primarily hydrolyze soluble substrates, as a candidate that accounted for some of the differences in hydrolytic activities. Heterologous expression confirmed that the GH12 protein identified by proteomics was active on crystalline cellulose and hydrolyzed cellulose by a random mechanism, in contrast to most cellulases that act on the crystalline polymer in a processive mechanism.

Original languageEnglish
Article numbere01106-16
JournalmBio
Volume7
Issue number4
DOIs
Publication statusPublished - Jul 1 2016
Externally publishedYes

Fingerprint

Glycoside Hydrolases
Cellulose
Proteomics
Hydrolysis
Proteins
Cellulases
Carbon Cycle
Biomass
Polymers
Enzymes
Microbial Consortia
Biofuels
Actinobacteria
Polysaccharides

ASJC Scopus subject areas

  • Microbiology
  • Virology

Cite this

Comparative community proteomics demonstrates the unexpected importance of actinobacterial glycoside hydrolase family 12 protein for crystalline cellulose hydrolysis. / Hiras, Jennifer; Wu, Yu Wei; Deng, Kai; Nicora, Carrie D.; Aldrich, Joshua T.; Frey, Dario; Kolinko, Sebastian; Robinson, Errol W.; Jacobs, Jon M.; Adams, Paul D.; Northen, Trent R.; Simmons, Blake A.; Singer, Steven W.

In: mBio, Vol. 7, No. 4, e01106-16, 01.07.2016.

Research output: Contribution to journalArticle

Hiras, J, Wu, YW, Deng, K, Nicora, CD, Aldrich, JT, Frey, D, Kolinko, S, Robinson, EW, Jacobs, JM, Adams, PD, Northen, TR, Simmons, BA & Singer, SW 2016, 'Comparative community proteomics demonstrates the unexpected importance of actinobacterial glycoside hydrolase family 12 protein for crystalline cellulose hydrolysis', mBio, vol. 7, no. 4, e01106-16. https://doi.org/10.1128/mBio.01106-16
Hiras, Jennifer ; Wu, Yu Wei ; Deng, Kai ; Nicora, Carrie D. ; Aldrich, Joshua T. ; Frey, Dario ; Kolinko, Sebastian ; Robinson, Errol W. ; Jacobs, Jon M. ; Adams, Paul D. ; Northen, Trent R. ; Simmons, Blake A. ; Singer, Steven W. / Comparative community proteomics demonstrates the unexpected importance of actinobacterial glycoside hydrolase family 12 protein for crystalline cellulose hydrolysis. In: mBio. 2016 ; Vol. 7, No. 4.
@article{49d0949081dc4a12bb7dd9d34af99a70,
title = "Comparative community proteomics demonstrates the unexpected importance of actinobacterial glycoside hydrolase family 12 protein for crystalline cellulose hydrolysis",
abstract = "Glycoside hydrolases (GHs) are key enzymes in the depolymerization of plant-derived cellulose, a process central to the global carbon cycle and the conversion of plant biomass to fuels and chemicals. A limited number of GH families hydrolyze crystalline cellulose, often by a processive mechanism along the cellulose chain. During cultivation of thermophilic cellulolytic microbial communities, substantial differences were observed in the crystalline cellulose saccharification activities of supernatants recovered from divergent lineages. Comparative community proteomics identified a set of cellulases from a population closely related to actinobacterium Thermobispora bispora that were highly abundant in the most active consortium. Among the cellulases from T. bispora, the abundance of a GH family 12 (GH12) protein correlated most closely with the changes in crystalline cellulose hydrolysis activity. This result was surprising since GH12 proteins have been predominantly characterized as enzymes active on soluble polysaccharide substrates. Heterologous expression and biochemical characterization of the suite of T. bispora hydrolytic cellulases confirmed that the GH12 protein possessed the highest activity on multiple crystalline cellulose substrates and demonstrated that it hydrolyzes cellulose chains by a predominantly random mechanism. This work suggests that the role of GH12 proteins in crystalline cellulose hydrolysis by cellulolytic microbes should be reconsidered. IMPORTANCE Cellulose is the most abundant organic polymer on earth, and its enzymatic hydrolysis is a key reaction in the global carbon cycle and the conversion of plant biomass to biofuels. The glycoside hydrolases that depolymerize crystalline cellulose have been primarily characterized from isolates. In this study, we demonstrate that adapting microbial consortia from compost to grow on crystalline cellulose generated communities whose soluble enzymes exhibit differential abilities to hydrolyze crystalline cellulose. Comparative proteomics of these communities identified a protein of glycoside hydrolase family 12 (GH12), a family of proteins previously observed to primarily hydrolyze soluble substrates, as a candidate that accounted for some of the differences in hydrolytic activities. Heterologous expression confirmed that the GH12 protein identified by proteomics was active on crystalline cellulose and hydrolyzed cellulose by a random mechanism, in contrast to most cellulases that act on the crystalline polymer in a processive mechanism.",
author = "Jennifer Hiras and Wu, {Yu Wei} and Kai Deng and Nicora, {Carrie D.} and Aldrich, {Joshua T.} and Dario Frey and Sebastian Kolinko and Robinson, {Errol W.} and Jacobs, {Jon M.} and Adams, {Paul D.} and Northen, {Trent R.} and Simmons, {Blake A.} and Singer, {Steven W.}",
year = "2016",
month = "7",
day = "1",
doi = "10.1128/mBio.01106-16",
language = "English",
volume = "7",
journal = "mBio",
issn = "2161-2129",
publisher = "American Society for Microbiology",
number = "4",

}

TY - JOUR

T1 - Comparative community proteomics demonstrates the unexpected importance of actinobacterial glycoside hydrolase family 12 protein for crystalline cellulose hydrolysis

AU - Hiras, Jennifer

AU - Wu, Yu Wei

AU - Deng, Kai

AU - Nicora, Carrie D.

AU - Aldrich, Joshua T.

AU - Frey, Dario

AU - Kolinko, Sebastian

AU - Robinson, Errol W.

AU - Jacobs, Jon M.

AU - Adams, Paul D.

AU - Northen, Trent R.

AU - Simmons, Blake A.

AU - Singer, Steven W.

PY - 2016/7/1

Y1 - 2016/7/1

N2 - Glycoside hydrolases (GHs) are key enzymes in the depolymerization of plant-derived cellulose, a process central to the global carbon cycle and the conversion of plant biomass to fuels and chemicals. A limited number of GH families hydrolyze crystalline cellulose, often by a processive mechanism along the cellulose chain. During cultivation of thermophilic cellulolytic microbial communities, substantial differences were observed in the crystalline cellulose saccharification activities of supernatants recovered from divergent lineages. Comparative community proteomics identified a set of cellulases from a population closely related to actinobacterium Thermobispora bispora that were highly abundant in the most active consortium. Among the cellulases from T. bispora, the abundance of a GH family 12 (GH12) protein correlated most closely with the changes in crystalline cellulose hydrolysis activity. This result was surprising since GH12 proteins have been predominantly characterized as enzymes active on soluble polysaccharide substrates. Heterologous expression and biochemical characterization of the suite of T. bispora hydrolytic cellulases confirmed that the GH12 protein possessed the highest activity on multiple crystalline cellulose substrates and demonstrated that it hydrolyzes cellulose chains by a predominantly random mechanism. This work suggests that the role of GH12 proteins in crystalline cellulose hydrolysis by cellulolytic microbes should be reconsidered. IMPORTANCE Cellulose is the most abundant organic polymer on earth, and its enzymatic hydrolysis is a key reaction in the global carbon cycle and the conversion of plant biomass to biofuels. The glycoside hydrolases that depolymerize crystalline cellulose have been primarily characterized from isolates. In this study, we demonstrate that adapting microbial consortia from compost to grow on crystalline cellulose generated communities whose soluble enzymes exhibit differential abilities to hydrolyze crystalline cellulose. Comparative proteomics of these communities identified a protein of glycoside hydrolase family 12 (GH12), a family of proteins previously observed to primarily hydrolyze soluble substrates, as a candidate that accounted for some of the differences in hydrolytic activities. Heterologous expression confirmed that the GH12 protein identified by proteomics was active on crystalline cellulose and hydrolyzed cellulose by a random mechanism, in contrast to most cellulases that act on the crystalline polymer in a processive mechanism.

AB - Glycoside hydrolases (GHs) are key enzymes in the depolymerization of plant-derived cellulose, a process central to the global carbon cycle and the conversion of plant biomass to fuels and chemicals. A limited number of GH families hydrolyze crystalline cellulose, often by a processive mechanism along the cellulose chain. During cultivation of thermophilic cellulolytic microbial communities, substantial differences were observed in the crystalline cellulose saccharification activities of supernatants recovered from divergent lineages. Comparative community proteomics identified a set of cellulases from a population closely related to actinobacterium Thermobispora bispora that were highly abundant in the most active consortium. Among the cellulases from T. bispora, the abundance of a GH family 12 (GH12) protein correlated most closely with the changes in crystalline cellulose hydrolysis activity. This result was surprising since GH12 proteins have been predominantly characterized as enzymes active on soluble polysaccharide substrates. Heterologous expression and biochemical characterization of the suite of T. bispora hydrolytic cellulases confirmed that the GH12 protein possessed the highest activity on multiple crystalline cellulose substrates and demonstrated that it hydrolyzes cellulose chains by a predominantly random mechanism. This work suggests that the role of GH12 proteins in crystalline cellulose hydrolysis by cellulolytic microbes should be reconsidered. IMPORTANCE Cellulose is the most abundant organic polymer on earth, and its enzymatic hydrolysis is a key reaction in the global carbon cycle and the conversion of plant biomass to biofuels. The glycoside hydrolases that depolymerize crystalline cellulose have been primarily characterized from isolates. In this study, we demonstrate that adapting microbial consortia from compost to grow on crystalline cellulose generated communities whose soluble enzymes exhibit differential abilities to hydrolyze crystalline cellulose. Comparative proteomics of these communities identified a protein of glycoside hydrolase family 12 (GH12), a family of proteins previously observed to primarily hydrolyze soluble substrates, as a candidate that accounted for some of the differences in hydrolytic activities. Heterologous expression confirmed that the GH12 protein identified by proteomics was active on crystalline cellulose and hydrolyzed cellulose by a random mechanism, in contrast to most cellulases that act on the crystalline polymer in a processive mechanism.

UR - http://www.scopus.com/inward/record.url?scp=84986634245&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84986634245&partnerID=8YFLogxK

U2 - 10.1128/mBio.01106-16

DO - 10.1128/mBio.01106-16

M3 - Article

AN - SCOPUS:84986634245

VL - 7

JO - mBio

JF - mBio

SN - 2161-2129

IS - 4

M1 - e01106-16

ER -