Solving the structure of a cellulose synthase Plant Conserved Region (PCR)

Ruston, PS; Olek, AT; Makowski, L; Badger, J; Steussy, CN,; Carpita, NC; andStauffacher, CV. The rice (Oryza sativa) Cellulose SynthaseA8 Plant-ConservedRegion is an anti-parallel coiled-coil located at the substrate entrance to thecatalytic core. Plant Phys. (2016) [10.1104/pp.16.00739]This work was performed at Purdue University, Northeastern University andArgonne National Laboratory

Scientific Achievement‐ We solved the crystal structure of one of two plant-

unique sequences in the catalytic domain of a cellulose synthase

Significance and Impact• Two antiparallel a -helices form a coiled-coil domain

linked by a large extended connector loop containing a

conserved trio of aromatic residues.

• The P-CR structure and molecular envelope were

modeled into the SAXS-derived catalytic core to

produce a detailed topological model of the CesA8

catalytic domain.

• The predicted position for the P-CR domain from the

molecular docking models places the coiled coil near

the entrance of the substrate UDP-glucose into the

active site.

• Our detailed topological model of the catalytic

monomeric domain of plant CesA fits via PCR

interactions into the trimeric structure determined after

renaturation of detergent soluble protein (Vandavasi et

al., 2015)

• However, these structures are unlikely to represent the

vivo structure due to clashes with the membrane

channels if present.

Fuels from Catalytic Hydrodeoxygenation of Lignin-Derived Oxygenates

Scientific Achievement: Complete removal ofoxygen from lignin model compound dihydroeugenol

Significance and Impact• Both Pt (hydrogenation) and Mo (dehydration) functions

are needed. PtMo catalyzes phenolic oxygen removal• Hydrogen pressure determines product; propyl benzene

at low pressure, propyl cyclohexane at high pressure• Direct pathway from lignin-derived molecules to fuels• Retains ring structure inherent in lignin

Sara L. Yohe, Harshavardhan J. Choudhari, Dhairya D.Mehta, Paul J. Dietrich, Michael D. Detwiler, Cem M.Akatay, Eric A. Stach, Jeffrey T. Miller, W. NicholasDelgass, Rakesh Agrawal, Fabio H. Ribeiro. High-pressure vapor-phase hydrodeoxygenation of lignin-derived oxygenates to hydrocarbons by a PtMobimetallic catalyst: Product selectivity, reactionpathway, and structural characterization. Journal ofCatalysis, 344, 535–552, (2016).[10.1016/j.jcat.2016.10.009]Work was performed at Purdue University

Research Details• Reaction pathway determined from

product distribution vs. conversion• PtMo alloy formation confirmed by

Scanning electron microscopy (STEM), x-ray absorption spectroscopy, and x-ray photoelectron microscopy

• Inert multiwalled carbon nanotubes used as support to facilitate electron-microscopy studies

• In addition to Pt and PtMo metal particles, Mo carbide, MoO2, and MoO3 phases were also present

Reaction pathwayHAADF STEM and

STEM-EELS of PtMocatalyst

OCH3

HO HO

HO

HO

OCH3

HO HO

H2O

CH4

H2O

H2OCH3OHPt

Pt Pt

Pt-Mo

Pt-MoPt

Pt

Pt

14.2 psi H2

97 psi H2342 psi H2

CH3OH

Scientific Achievement‐ Native secretory ferritins exist only in insects and worms, but

not in plants. ‐ We generated transgenic Arabidopsis that delivered ferritin

to cell wall region (FerEX) during plant growth.‐ FerEX biomass had 20‐35% higher pretreatability and

digestibility than the control and ~10% higher than intracellularly expressed ferritin plants (FerIN).

Significance and Impact‐ It effectively eliminates the current approach of soaking iron‐

containing acid solutions onto milled biomass prior to pretreatment, which is time‐consuming and subject to diffusion limitations. Research Details

Directed plant cell-wall accumulation of iron: Embedding co-catalyst for efficient biomass conversion

Glucose and xylose release increases 20-35%

during pretreatment

Joint efforts with USDA FPL/APS Argonne: X-rayfluorescence microscopy (XFM) maps for Fe in stemcross-sections

FerEX transgenic is hyper iron-

accumulated, and 12% taller and

18% more dry mass than control

Transgenic poplar >60% more

mass

Lin, Chien-Yuan; Jakes, Joseph E.; Donohoe, Bryon S.; Ciesielski, Peter N.; Yang, Haibing;Gleber, Sophie-Charlotte; Vogt, Stefan; Ding, Shi-You; Peer, Wendy A.; Murphy, Angus S.;McCann, Maureen C.; Himmel, Michael E.; Tucker Melvin P.; and Wei, Hui. Directed plant cellwall accumulation of iron: Embedding co-catalyst for efficient biomass conversion.Biotechnology for Biofuels 9, 225. [10.1186/s13068-016-0639-2]Work was performed at NREL, USDA, Argonne Nat.Lab, and Purdue University

Enhanced Acid-Catalyzed Biomass Conversion to HMF Following COSLIF Pretreatment

Scientific Achievement‐ Concentrated phosphoric acid decrystallization of

cellulose enhances acid-catalyzed conversion of cellulose directly to HMF

Significance and Impact‐ Maleic acid in combination with aluminum chloride has

been shown to be effective for isomerizing glucose to fructose (Al3+) and dehydrating fructose to HMF (maleic acid).

‐ Acid hydrolysis of cellulose is hampered by its crystallinity.‐ Concentrated phosphoric acid (COSLIF) decrystallizes

cellulose without changing the biomass composition.‐ Decrystallized cellulose (Avicel) is more readily converted

to HMF and at higher yields.‐ The impact of COSLIF is less dramatic for intact biomass

Hewetson, B. H.; Zhang, X., Mosier, N.S. Enhanced Acid-Catalyzed BiomassConversion to Hydroxymethylfurfural Following Cellulose Solvent- and OrganicSolvent-Based Lignocellulosic Fractionation Pretreatment, Energy & Fuels, , 30,9975–9977 (2016). [10.1021/acs.energyfuels.6b01910]Work was performed at Purdue University

Research Details‐ COSLIF decrystallization used 75% phosphoric acid, 50 C, 60

minutes followed by ethanol precipitation of dissolved cellulose.

‐ Conversion of 2.5 wt% suspension of cellulose/biomass in equimolar concentrations of maleic acid and AlCl3 (25 mM), 180 C for up to 40 minutes.

Yields (percentage of theoretical) for cellulose-derived products with and without COSLIF treatment.

MSn Based on Collision-Activated Dissociation (CAD) for Identification of Compounds Related to Lignin

Scientific Achievement‐ Development of a method to identify lignin conversion products

Significance and Impact‐ The primary obstacle to the use of lignin as a fuel is its high

oxygen content. Conversion methods have been developed to address this issue. Evaluation of their value requires methods for the identification of lignin conversion products. This is difficult due to the complexity of the product mixtures.

‐ MS6/CAD experiments and quantum chemical calculations allowed the identification of many diagnostic reactions and delineation of their mechanisms to ensure predictable behavior

‐ Above information was utilized to identify the presence of specific functionalities and their combinations in molecules in a complex organosolv lignin sample

Marcum, Christopher L.; Tiffany M. Jarrell, Hanyu Zhu, Benjamin C. Owen, Laura J. Haupert,Hilkka I. Kenttämaa A Fundamental Tandem Mass Spectrometry Study of the Collision‐Activated Dissociation of Small Deprotonated Molecules Related to Lignin, ChemSusChem.[10.1002/cssc.201600678]; Work was performed at NREL and Purdue University

Research Details‐ The CAD pathways and mechanisms of 34 deprotonated

lignin related model compounds were explored experimentally (up to MS6) and computationally (M06‐2X/6‐311++G(d,p)//M06‐2X/6‐311++G(d,p)).

‐ The compounds were ionized using negative‐ion mode electrospray ionization doped with NaOH in order to produce abundant intact deprotonated molecules(method developed by us for C3Bio).

Figure 1. Top: MS7 experiment. Bottom: HPLC/MS6 analysis of an organosolv lignin sample obtained from switchgrass.

Heterogeneous Diels-Alder catalysis for biomass-derived aromatic compounds

Scientific Achievement‐ Published technical review article on routes to upgrade

biomass to aromatic monomers via Diels-Alder catalysis

Significance and Impact‐ Sugar and lignin‐derived monomers can be utilized for

producing aromatic monomers via Diels‐Alder chemistry‐ Multifunctional heterogeneous catalyst can be tailored

for both acidity and dehydrogenation activity‐ Route offers access to direct replacement commodity

aromatics, as well as novel functional alternatives‐ Article highlighted for the inside front cover image in the

journal Green Chemistry

Settle, A.E., Berstis, L., Rorrer, N.A., Roman-Leshkóv, Y., Beckham,G.T., Richards, R.M., Vardon, D.R., 2017. Heterogeneous Diels-Aldercatalysis for biomass-derived aromatic compounds. Green Chemistry,19, 3468-3492. [10.1039/C7GC00992E]Work was performed at NREL

Research Details‐ Diels‐Alder (DA) catalysis review addresses:

‐ Reaction mechanism aspects from both an experimental and computational perspective

‐ Tailored Brønsted‐Lewis acid design strategies for heterogeneous catalysts (e.g., zeolites and POMs)

‐ Novel aromatic monomers with chemical functionality that is difficult to access from petroleum

Rhamnogalacturonan I, xylan and lignin are determinants of cell-cell adhesion in poplar wood

Scientific Achievement‐ We identified a new function for the pecticpolymer, rhamnogalacturonan I (RG-I), in cell-cell adhesion in poplar.

Significance and Impact‐ Cell‐cell adhesion occurs at wall domains of distinct mesoscale

architectures but the molecular bases of this property have not been studied in bioenergy‐relevant species.

‐ Complete cell separation can be effected by removing lignin, xylan, and RG‐1.

‐ Genetic control of RG‐1 content in poplar cell walls by expression of an RG‐lyase activity modulates cell‐cell adhesion and catalyst accessibility.

‐ Genetic control of cell‐cell adhesion is one strategy to reduce energy inputs in biomass comminution.

Work was performed at Purdue University

Research Details‐ An Arabidopsis RG‐lyase was expressed in poplar and multiple

independent transgenic lines recovered.‐ Numbers of single cells released from particles are positively

correlated with transcript abundance and RG‐lyase activity.‐ High‐expressing lines show increased yield of sugars and accessibility

to catalysts with or without prior extraction of lignin and xylan.

WT RGIL6-2 RGIL6-7 RGIL6-34

Time (h)