木质纤维素的微生物降解

木质纤维素广泛存在于自然界中,因结构复杂,其高效降解需要多种微生物的协同互作,由于参与木质纤维素降解的微生物种类繁多,其协同降解机理尚不完全明确。随着微生物分子生物学和组学技术的快速发展,将为微生物协同降解木质纤维素机制的研究提供新的方法和思路。笔者前期研究发现,细菌复合菌系在50℃下表现出强大的木质纤维素降解能力,菌系由可分离培养和暂时不可分离培养细菌组成,但是可分离培养细菌没有降解能力。通过宏基因组和宏转录组研究表明,与木质纤维素降解相关的某些基因表达量发生显著变化,通过组学方法有可能更加深入解释微生物协同降解木质纤维素的微生物学和酶学机理。文中从酶、纯培养菌株和复合菌群三个方面综述了木质纤维素微生物降解研究进展,着重介绍了组学技术在解析复合菌群作用机理方面的现状和应用前景,以期为探索微生物群落协同降解木质纤维素的机理提供借鉴。     

Microbial degradation of lignin

The transformations of lignin that occur during its biodegradation are complex and incompletely understood. Certain fungi of the white-rot group, and possibly other fungi and bacteria, completely decompose lignin to carbon dioxide and water. Other fungi and bacteria apparently degrade lignin incompletely. Differences in lignin-degrading abilities observed for different organisms may result from differences in the completeness of their ligninolytic enzyme systems. Not all lignin components may be attacked by a particular organism. Alternatively, different organisms may differ in their basic mechanisms of attack on lignin. The basic pathways of lignin degradation have been elucidated only for certain representatives of the white-and brown-rot fungi. Although it is known that each of the principal structural components of lignin is attacked by other fungi and bacteria, the biochemistry of that attack has not been elucidated. Work with low molecular weight lignin models has provided only limited information on possible pathways of lignin degradation by microorganisms. There is little evidence to suggest a correlation between abilities to degrade single-ring aromatic or lignin model compounds and the ability to degrade polymeric lignin. More evidence has come from analysis of spent culture media for lignin breakdown products and from comparative chemical analyses of sound lignins versus decayed lignin residues. Accumulated evidence with the most thoroughly studied white-rot fungi suggests that with these fungi lignin degradation proceeds by way of extracellular mixed-function oxygenases and dioxygenases, which catalyse demethylations, hydroxylations and ring-fission reactions within a largely intact polymer, concomitant with some release of low molecular weight lignin fragments. There are also apparent relationships between lignin, carbohydrate and nitrogen metabolism for some organisms, but the relationships may vary from one organism to another. Although research is now mostly at a basic level, industrial applications may result from lignin degradation research. Considerable potential exists for the development of bioconversions which might produce low molecular weight chemicals from waste lignins, and thereby reduce our dependence on petroleum as a source of these chemicals. Alternatively, such bioconversions might produce chemically altered forms of polymeric lignin that may be valuable industrially.     

微生物法降解木质素的研究进展

生物质是代替石化资源生产能源和化学品的关键资源,木质素作为植物细胞壁的主要成分已经在很多行业中得到了广泛的应用。然而,由于木质素结构复杂且难以降解,成为生物质资源利用的最大障碍,因此,去除或者降解木质素是利用细胞壁中其他成分的关键步骤。许多行业使用有害化学物质降解木质素,严重危害了生态环境,自然界中木质素经常被包括真菌和细菌在内的微生物降解,因此,研究微生物降解木质素的机制为解决这一问题提供了可能性。本文讨论了木质素的化学组成成分,重点讨论了自然界降解木质素的微生物种类及其降解机制,包括各种真菌和细菌的木质素降解活性,描述了由各种微生物特别是白腐真菌、褐腐真菌和细菌产生的木质素降解酶,并展望了今后木质素生物降解的研究和应用的可能方向。     

Factors affecting lignin degradation in lignocellulose by Phanerochaete chrysosporium

Cultural conditions affecting lignin degradation by Phanerochaete chrysosporium in various lignocellulosic materials were studied in comparison to an isolated lignin preparation. With shallow mycelial cultures, the degradation of lignin in wood proceeded more slowly in a 100% O₂-atmosphere than in an air atmosphere, indicating that pure oxygen was toxic to the fungus. The organism was able to degrade lignin efficiently even under 30% CO₂ and 10% O₂ concentrations. Evolution of ¹⁴CO₂ from labelled lignocellulosic materials was shown not to be representative of total lignin degradation. Addition of glucose to the culture did not affect lignin degradation measured by ¹⁴CO₂ evolution, whereas lignin degradation measured by Klason lignin method stopped completely (poplar) or slowed considerably (straw). Due to partial depolymerization of lignin to soluble products, measuring only the evolution of ¹⁴CO₂ results in an underestimation of the total amount of lignin bioaltered. The soluble products from all of the tested lignocellulosic materials and from the isolated lignin had an average molecular weight of about 1,000 and the products could be further fractionated by ion exchange chromatography. The relative amount of these products could be varied from 15 to 45% from the original lignin.     

Microbial community succession and lignocellulose degradation during agricultural waste composting

The changes of microbial community during agricultural waste composting were successfully studied by quinone profiles. Mesophilic bacteria indicated by MK-7 and mesophilic fungi containing Q-9 as major quinone were predominant and seemed to be important during the initial stage of composting. Actinobacteria indicated by a series of partially saturated and long-chain menaquinones were preponderant during the thermophilic period. While Actinobacteria, fungi and some bacteria, especially those microbes containing MK-7(H4) found in Gram-positive bacteria with a low G+C content or Actinobacteria were found cooperate during the latter maturating period. Since lignocellulsoe is abundant in the agricultural wastes and its degradation is essential for the operation of composting, it’s important to establish the correlation between the quinone profiles changes and lignocellulose degradation. The microbes containing Q-9 or Q-10(H2) as major quinone were found to be the most important hemicellulose and cellulose degrading microorganisms during composting. While the microorganisms containing Q-9(H2) as major quinone and many thermophilic Actinobacteria were believed to be responsible for lignin degradation during agricultural waste composting.     

Microbial utilization of lignin: available biotechnologies for its degradation and valorization

Lignocellulosic biomasses, either from non-edible plants or from agricultural residues, stock biomacromolecules that can be processed to produce both energy and bioproducts. Therefore, they become major candidates to replace petroleum as the main source of energy. However, to shift the fossil-based economy to a bio-based one, it is imperative to develop robust biotechnologies to efficiently convert lignocellulosic streams in power and platform chemicals. Although most of the biomass processing facilities use celluloses and hemicelluloses to produce bioethanol and paper, there is no consolidated bioprocess to produce valuable compounds out of lignin at industrial scale available currently. Usually, lignin is burned to provide heat or it remains as a by-product in different streams, thus arising environmental concerns. In this way, the biorefinery concept is not extended to completion. Due to Nature offers an arsenal of biotechnological tools through microorganisms to accomplish lignin valorization or degradation, an increasing number of projects dealing with these tasks have been described recently. In this review, outstanding reports over the last 6 years are described, comprising the microbial utilization of lignin to produce a variety of valuable compounds as well as to diminish its ecological impact. Furthermore, perspectives on these topics are given.     

Microbial degradation of dehydrodiconiferyl alcohol,a lignin substructure model

Bacteria, yeasts, and molds which grew in a medium containing a synthetic lignin — a dehydrogenation polymer (DHP) of coniferyl alcohol — as a sole carbon source, were isolated from soil. One fungus, Fusarium solani M-13-1, was found to degrade the DHP most vigorously among the isolated organisms. It was shake-cultured in a medium containing dehydrodiconiferyl alcohol (DHCA) (I), an important lignin model compound, and the following six metabolic products were isolated and identified: 1) Phenylcoumaran--aldehydic (II) and -carboxylic compounds, 2) phenylcoumaran--aldehydic compound (IV), formed by release of a 2-carbon fragment from the phenylcoumaran--carboxylic compound, 3) 5-acetylvanillyl alcohol (V), formed by cleavage of the coumaran ring and reduction of the -aldehyde group, 4) 5-carboxyvanillyl alcohol (VI), formed by subsequent oxidation of the acetyl group, and 5) the -ether of DHCA (VII), considered to be a by-product. A degradation pathway for DHCA was proposed on the basis of these metabolic products.Non-Standard Abbreviations DHP dehydrogenation polymer - DHCA dehydrodiconiferyl alcohol - DDQ dichlorodicyano-p-benzoquinone - DDHQ dichlorodicyano-p-hydroquinone - Ar aromatic - TLC thin layer chromatography - GC-MS gas chromatography-mass spectrometry     

Immunological characterization of lignocellulose degradation

Polyclonal antibodies produced to the brown-rot fungus Poria placenta have been used with fluorescence microscopy to detect fungal hyphae colonizing wood. In addition, an enzyme-linked immunosorbent assay (ELISA) has been used to detect and quantify the extent of fungal decay in wood. ELISA values correlated with percent weight loss associated with fungal degradation and the assay was able to detect fungal colonization ten days after inoculation in solid wood. Monoclonal antibodies to extracellular metabolites of P. placenta have been produced and are being characterized. Monoclonal antibodies have also been produced to native and partially deglycosylated Mn2 peroxidase. The use of immunological probes to monitor and characterize the decay process is discussed.     

Anaerobic biodegradation of the lignin and polysaccharide components of lignocellulose and synthetic lignin by sediment microflora

[This corrects the article on p. 998 in vol. 47.].     

Biotechnology in the degradation and utilization of lignocellulose

Lignocellulose is the predominant renewable resource. It uses include fuel, as the feedstock for the pulp and paper industry, and for animal nutrition. It also constitutes a large proportion of agricultural and urban waste. Biotechnology has roles in its efficient production and utilisation. The types of lignin substrates available for study of lignin biodegradation are described. The white rot fungus Phanerochaete chrysosporium is the archetypal system for the study of lignocellulose degradation, since it mineralises lignin and degrades both cellulose and hemicellulose. The salient features of the P. chrysosporium system are described. The lignin peroxidases are a family of proteins, and it is shown that expression of their genes is differential. P. chrysosporium is heterokaryotic with two gene equivalents that have abundant RFLPs. A set of basidiospore-derived strains with genetic compositions defined by such RFLPs provided the potential basis for a strain improvement programme for lignin degradation. However, analysis of this system using radiolabelled synthetic lignin (DHP) as the substrate confirmed previous evidence that both the substrate and the fungal cultures displayed much variation, so that it was difficult to quantify performance for this property. The cellobiohydrolase I enzymes are also coded for by a family of genes, and evidence is also presented for allelic variants, for differential expression and for differential splicing. In contrast, the cellobiohydrolase II function is encoded at a unique genetic locus. Approaches to an homologous integrative transformation system are discussed. Some actinomycete bacteria represent an alternative system for lignin solubilisation in which strains differ in their spectra of activities on lignocellulose substrates. The xylanase system of Streptomyces cyaneus is shown to include three enzymes, two of which are inducible by xylan. A novel assay method was developed and used to demonstrate that the third is constitutive and also non-repressible by glucose. It is proposed that this acts as a sensor for xylans in the environment that can yield breakdown products that are taken up and can then act as inducers of the other two enzymes. The studies on microbial lignocellulose degradation from different laboratories have allowed the formulation of specific biotechnological goals, and some of the problems and opportunities in this area are identified.     

On the bacterial degradation of lignin

Summary Several bacteria which can degrade numerous phenols with structural relationships to lignin were tested for their ability to degrade lignin. The biodegradation with all the tested bacteria was poor. The method of lignin extraction, presence of glucose as cosubstrate and changes in the nitrogen source of the medium did not affect the extent of lignin degradation. The poor degradation does not seem to be influenced by medium composition and culture condition but is more probably due to the inability of the tested bacteria to degrade lignin to any considerable extent.     

Bacteria and lignin degradation

Lignin is both the most abundant aromatic (phenolic) polymer and the second most abundant raw material.It is degraded and modified by bacteria in the natural world,and bacteria seem to play a leading role in decomposing lignin in aquatic ecosystems.Lignin-degrading bacteria approach the polymer by mechanisms such as tunneling,erosion,and cavitation.With the advantages of immense environmental adaptability and biochemical versatility,bacteria deserve to be studied for their ligninolytic potential.     

Bacteria and lignin degradation

Lignin is both the most abundant aromatic (phenolic) polymer and the second most abundant raw material. It is degraded and modified by bacteria in the natural world, and bacteria seem to play a leading role in decomposing lignin in aquatic ecosystems. Lignin-degrading bacteria approach the polymer by mechanisms such as tunneling, erosion, and cavitation. With the advantages of immense environmental adaptability and biochemical versatility, bacteria deserve to be studied for their ligninolytic potential.     

Acid-precipitable polymeric lignin from hardwood lignocellulose: production by a Streptomyces sp.

A Streptomyces sp. isolate, from decayed wood shavings, solubilized lignocellulose (LC) and lignin of Pinus radiata, producing about 50 mg acid-precipitable polymeric lignin per g LC. The product was poor in protein and carbohydrates and contained mainly vanillin, guaicol, vanillic and ferulic acids. Hardwood LC is thus suitable for producing APPL as a phenolic chemical feedstock.V.M. Kaluskar is with the Department of Microbiology, J and J Science College, Nadiad 387001, Gujarat, India. B.P. Kapadanis is with the Department of Microbiology, School of Sciences, University of Pune, Ganesh Khind, Pune-41107, Maharashtra, India. M.J. Penninckx is with the Unit of Microbial Physiology and Ecology, Free University of Brussels, c/o IPB 642, rue Engeland, B-1180, Brussels, Belgium     

细菌降解木质素的研究进展

木质素是自然界最丰富的芳香化合物,其分解与陆地上碳循环密切相关。提取木质纤维素中的葡萄糖使其转化成乙醇,是生产第二代生物能源的关键步骤。但是由于木质素是一种非常稳定的化合物,难以降解是实现生物乙醇转化的主要屏障,因此关于木质素的生物降解研究具有非常重要的意义。真菌降解木质素的研究已经深入的进行了多年,并取得丰富的成果,但是关于细菌降解木质素的研究还处在初级阶段。由于广泛的生长条件和良好的环境适应能力,细菌在木质素降解方面深受研究人员的关注。本文通过总结前人的研究成果,讨论了木质素的降解机制、代谢途径及细菌降解木质素的工业应用前景,同时还展望了分子生物学及生物信息学在木质素降解方面的应用前景。     

Microbial tolerance engineering toward biochemical production: from lignocellulose to products

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Enzymatic grafting of functional molecules to the lignin model dibenzodioxocin and lignocellulose material

Laccase-mediated grafting of functional molecules presents an eco-friendly approach to functionalize lignocellulose materials. In this study functional molecules in the form of reactive phenolic amines, hydrophobicity enhancing fluorophenols and selected wood preservatives, were for the first time successfully coupled onto the lignin model compound dibenzodioxocin (Db) as demonstrated by HPLC-MS analysis. A 1:1-coupling was demonstrated for various combinations including Db and tyramine (m/z 620.5), Db and 3-O-methyldopamine (m/z 650.5), Db and 4-hydroxy-3-methoxybenzylamine (m/z 636.5), Db and 4-fluoro-2-methylphenol (m/z 609.5), and Db and 2-phenylphenol (m/z 653.5). Fungal laccases from Trametes hirsuta and T. villosa were more efficient in mediating the coupling of tyramine to dibenzodioxocin and beech (Fagus sylvatica) wood than a Bacillus sp. laccase with lower redox potential. This work presents for the first time a model for functionalizing of lignocellulose using the lignin model dibenzodioxocin.