Renewable and Sustainable Energy Reviews 105 (2019) 349–362 Contents lists available at ScienceDirect Renewable and Sustainable Energy Reviews journal homepage: www.elsevier.com/locate/rser Identifying and creating pathways to improve biological lignin valorization T ⁎ Zhi-Hua Liua,b,1, Rosemary K. Lec,1, Matyas Kosad, Bin Yange, Joshua Yuana,b, , ⁎⁎ Arthur J. Ragauskasc,f,g, a Synthetic and Systems Biology Innovation Hub (SSBiH), Texas A&M University, College Station, TX 77843, United States b Department of Plant Pathology and Microbiology, Texas A&M University, College Station, TX 77843, United States c Department of Chemical & Biomolecular Engineering, University of Tennessee Knoxville, Knoxville, TN 37996, United States d Fortress Advanced Bioproducts Inc., Vancouver, BC, Canada V6T 1W5 e Bioproducts, Sciences, and Engineering Laboratory, Department of Biological Systems Engineering, Washington State University, Richland, WA 99354, United States f Joint Institute for Biological Sciences, Biosciences Division, Oak Ridge National Laboratory, Oak Ridge, TN 37831, United States g Center for Renewable Carbon, Department of Forestry, Wildlife, and Fisheries, University Tennessee, Knoxville, TN 37996, United States ARTICLE INFO ABSTRACT Keywords: Biological lignin valorization to fuels and value-added chemicals enables sustainable and economic biorefineries. Biological lignin valorization While significant progress has been made, several major challenges arose due to high recalcitrance and het- Ligninolytic microorganism erogeneity of lignin, which needs to be addressed to improve lignin processing. This work provides an overview Bioprospecting of biological lignin conversion and its regulation from a metabolic engineering and systems biology viewpoint. Metabolic engineering Biological lignin valorization includes three stages: lignin depolymerization, aromatics degradation, and target Systems biology product biosynthesis. Ligninolytic microorganisms have an extensive enzymatic toolbox to break down the lignin Synthetic biology Techno-economic analysis and convert heterogeneous lignin derivatives to central intermediates, such as protocatechuate or catechol, through a peripheral pathway. These intermediates undergo ring cleavage via the β-ketoadipate pathway and are ultimately transformed into metabolites by yielding acetyl-CoA for internal product biosynthesis, such as tria- cylglycerols, polyhydroxyalkanoates, etc. Bioprospecting will expand the knowledge base of ligninolytic mi- crobial communities, strains, and enzymes to facilitate the understanding of aromatics metabolism. Systems biology analyses achieve an understanding of molecular and systems-level degradation mechanisms and meta- bolic pathways of lignin and aromatics. By identifying these mechanisms, synthetic biology provides promising approaches to create the lignin conversion pathways and engineer ligninolytic strains suitable as potential hosts for lignin conversion. Techno-economic analysis of biological lignin upgrading to coproducts in biorefineries will guide the implementation of lignin valorization by mitigating technical risk for scale-up and improving the profitability of biorefinery. By improving the understanding of biological lignin valorization, it should be pos- sible to create biological lignin valorization route to effectively produce value-added products from lignin. 1. Introduction considered to be suitable for the production of biofuels as well as bulk and fine chemicals, potentially addressing energy and environmental Demand for renewable resource based fuels and chemicals is on the concerns [1,3]. With the intensive development of biorefineries for rise due to the dramatic increase in greenhouse gas emissions [1,2]. The fuels and chemicals from carbohydrates, the amount of lignin-rich re- lignin polymer in plant biomass is the most abundant renewable source sidues has increased, warranting new strategies for upgrading lignin of aromatic carbon and the second most abundant terrestrial polymer [2,4–6]. For instance, in the cellulosic ethanol and pulp and paper in- after cellulose. Aromatic compounds derived from lignin have been dustries, about 112 million tons of lignin are annually produced as Abbreviations: CDW, Cell dry weight; DyP, Dye peroxidase; LiP, Lignin peroxidase; MnP, Manganese peroxidase; mcl-PHAs, Medium chain-length poly- hydroxyalkanoates; O2-KL, Oxygen pretreatment Kraft lignin; PAHs, Polycyclic aromatic hydrocarbons; PHA, Polyhydroxyalkanoates; TAG, Triacylglycerol; TCA, Tricarboxylic acid; TEA, Techno-economic analysis; US-EOL, Ultrasonicated ethanol organosolv lignin; WT, Wild type ⁎ Corresponding author at: Synthetic and Systems Biology Innovation Hub (SSBiH), Texas A&M University, College Station, TX 77843, United States. ⁎⁎ Corresponding author at: Department of Chemical & Biomolecular Engineering, University of Tennessee Knoxville, Knoxville, TN 37996, United States. E-mail addresses: [email protected] (Z.-H. Liu), [email protected] (R.K. Le), [email protected] (M. Kosa), [email protected] (B. Yang), [email protected] (J. Yuan), [email protected] (A.J. Ragauskas). 1 These authors contributed equally to this work. https://doi.org/10.1016/j.rser.2019.02.009 Received 3 June 2018; Received in revised form 18 January 2019; Accepted 12 February 2019 Available online 16 February 2019 1364-0321/ © 2019 Elsevier Ltd. All rights reserved. Z.-H. Liu, et al. Renewable and Sustainable Energy Reviews 105 (2019) 349–362 Fig. 1. Schematic structure of lignin biopolymer in plant biomass. inexpensive and readily available residue in the United States alone [7]. exactly controlled to synthesize the desired product in microorganisms. Recently, lignin valorization has attracted more attention and been Among available state-of-the-art technologies, biological lignin va- considered in plant biomass upgrading strategies for economic and lorization that involved lignin depolymerization and conversion by sustainable biorefineries. Even so, the commercial application of lignin microorganisms holds great potential. A desirable lignin bio-depoly- is limited to only a few examples (e.g. vanillin and lignosulfonates) merization protocol would be a series of reactions with high reaction because of the challenging approaches for lignin depolymerization. rates that also allow for bioconversion to value-added compounds. Lignin is biosynthesized in oxidative coupling reactions using cin- Despite significant progress, biological lignin valorization is still highly namyl alcohol derivatives: p-coumaryl alcohol, coniferyl alcohol, and challenging with low levels of useful products generated. Several major sinapyl alcohol (Fig. 1). It is an arguably random three-dimensional issues need to be addressed to further improve biological lignin valor- biopolymer composing of phenylpropanoid units linked together by ization. Herein this paper overviews the biological lignin biorefinery various linkages, including β-O-4-aryl ether, β-5-phenylcoumaran, 5–5- from a metabolic engineering and systems biology viewpoint, and biphenyl, β-β-resinol and 4-O-5-diaryl ether linkage [8–10]. β-O-4-aryl points out future prospects for microbial production of lignin-derived ether is the most abundant linkage in lignin polymer [3]. Lignin carries biofuels and chemical. a variety of functional groups, for example, aliphatic hydroxyl, car- bonyl, phenolic hydroxyl, methoxyl, and benzyl alcohol groups, which 2. Biological processing of lignin by ligninolytic microorganisms determine the polarity and quality of both native and technical lignin. These chemistries are responsible for its heterogeneity and high re- 2.1. Ligninolytic microorganisms calcitrance, imposing a unique challenge for its depolymerization. Numerous studies have been conducted in the past few decades into Biological processing of lignin has emerged as a potentially effective valorizing lignin, in which the resulting monomers and oligmers from approach to gain lignin-derived products. The benefits of using biolo- lignin depolymerization by biological, thermal, or chemical pretreat- gical processing include the ability to use renewable carbon sources as – ments are converted to fuels and chemicals [6,11 15]. Most pretreat- feedstock to sustain cellular growth for generating various inter- ment approaches for lignin depolymerization produce a heterogeneous mediates and the ability for target molecules to be expressed with a array of aromatic species from lignin, depending on the feedstock types specificity that surpasses synthetic chemistry [25–28]. The capability of – used and the pretreatment approaches employed [16 18]. A hetero- microorganisms to selectively create target chemicals with special ffi geneous mixture of aromatic compounds may be su cient to produce features can be harnessed to generate new molecules for unique bio- fuels; however, high purity of aromatic compounds is necessary for the fuels or chemicals. fi production of ne chemicals [11,12,17,19]. Therefore, heterogeneity of Both fungi and bacteria have been found capable of degrading lignin lignin or aromatics generated from these depolymerization processes in vitro and/or in vivo with multiple enzymes and associated small fi presents a major obstacle for upgrading lignin to ne chemicals. molecule co-factors. The degradation mechanisms of lignin and aro- Research of biological lignin processing has been ramped up and matics have also been studied in different natural biomass utilization screening of microorganisms in ligninolytic environments has led to the systems [23,29].