青霉生产木质纤维素降解酶系的研究进展

木质纤维素降解酶系的高效生产是实现植物生物质大规模生物炼制的重要支撑。就地生产木质纤维素降解酶,有助于降低其使用成本,提高技术经济效益。青霉是自然界常见的木质纤维素降解真菌,可以合成分泌种类多样、组分齐全的木质纤维素降解酶系,已被应用于纤维素酶制剂的工业生产。文中从就地生产降解酶,为木质纤维素生物炼制构建“糖平台”的角度,综述了青霉木质纤维素降解酶系的性质、菌株遗传改造及发酵工艺的研究进展。     

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。     

降解纤维素的“超分子机器”研究进展

综述了目前关于纤维小体组装模式、纤维小体结构多样性及人工设计纤维小体等方面的研究进展.纤维小体是某些厌氧菌产生的由多个亚基共同组装而成的大分子机器,是致力于组织、协调多种酶组分协同高效催化降解木质纤维素的胞外蛋白质复合体.纤维小体是厌氧微生物水解纤维素的主体,具有非常高效的打破结晶纤维素的结晶结构和降解纤维素链的作用.纤维小体对木质纤维素降解的高效性来自于其自发组装而成的复杂的高级结构,其结构的复杂性因不同的厌氧微生物而有所不同.     

纤维素乙醇统合加工过程中的酵母多酶共展示体系研究进展

利用统合生物加工过程(Consolidated bioprocessing,CBP)生产纤维素乙醇是目前国内外的研究热点。CBP需要一种“集成化”微生物,既能生产水解木质纤维素的多种酶类又能利用水解木质纤维素产生的糖类发酵产乙醇。以酿酒酵母表面展示技术为依托,建立CBP菌株多酶共展示体系的研究主要分为以下两个方向：一是直接将纤维素酶展示在细胞表面,即非复合型纤维素酶体系;另一种是通过表面展示纤维小体(Cellulosome)将纤维素酶间接地锚定在细胞表面,即复合型纤维素酶体系,本文主要从以上两个方向阐述了近几年对于纤维素乙醇生物统合加工过程的研究进展。因纤维小体对纤维素的降解能力比非复合型纤维素酶体系更强,所以其在酿酒酵母细胞表面的组装研究受到越来越多的关注,为了更深入透彻地了解纤维小体的酵母展示技术,文中对纤维小体的结构与功能及其在纤维素乙醇发酵中的应用研究进行重点论述,并对该领域的发展方向进行展望。     

运动发酵单胞菌在生物炼制中的研究进展

以木质纤维素生物质为原料的生物炼制技术已成为全球研发的热点和难点。欧盟国家和美国的中长期生物质能源发展路线图中均将木质纤维素生物炼制技术作为重要目标,但是目前整体水平尚处于中试阶段。我国的纤维素类生物质原料非常丰富,将其转化成燃料乙醇及生物基础化学品等具有较大的潜力,但当前要想实现商业化生产,还面临着很多瓶颈问题亟待解决。缺乏能够同时高效利用纤维素类水解物的发酵菌株,已成为纤维素生物质高效与高值转化的关键制约因素。运动发酵单胞菌是目前唯一一种通过ED途径兼性厌氧发酵葡萄糖的微生物,其独特的代谢途径使其成为构建产乙醇工程菌的优选宿主之一;同时由于该菌具有较高的糖利用效率等优点,也是其他生物基化学品生产的重要候选平台微生物,如山梨醇、葡萄糖酸、丁二酸和异丁醇等。本文从该菌的研究历程、分子生物学基础、菌种改良及该菌在生物能源及生物基化学品等生物炼制体系中的应用研究角度进行了综述,并提出该菌可作为纤维素生物质生物炼制系统的新的重要平台微生物。     

酿酒酵母人造纤维小体的研究进展简

纤维素乙醇的统合生物加工过程（consolidated bioprocessing,CBP）是将（半）纤维素酶生产、纤维素水解和乙醇发酵过程组合,通过一种微生物完成的生物加工过程。 CBP有利于降低生物转化过程的成本,受到研究者的普遍关注。酿酒酵母（ Saccharomyces cerevisiae）作为传统的乙醇生产菌株,是极具潜力的CBP底盘细胞。纤维小体是某些厌氧微生物细胞表面由纤维素酶系与支架蛋白组成的大分子复合物,它能高效降解木质纤维,在酿酒酵母表面展示纤维小体已成为构建CBP细胞的研究热点。笔者综述了人造纤维小体在酿酒酵母细胞表面展示组装的研究进展,重点阐述了纤维小体各元件的设计和改造,并针对酿酒酵母分泌途径的改造,提出提高人造纤维小体分泌组装的可能性策略。     

筛选微生物降解木质纤维素的研究进展

木质纤维素资源是自然界中含量丰富的可再生资源,利用微生物降解木质纤维素是一种重要的策略。在综合国内外对木质纤维素降解微生物的筛选方法和研究策略的基础上,从单一菌株、复合微生物菌系和组学技术三个方面对筛选微生物降解木质纤维素进行了总结和分析,阐述了各个策略的优势特点和应用价值,即单一菌株易于培养但降解能力较低,复合菌系降解能力强但传代稳定性较差,组学技术能够更好的解释微生物降解木质纤维素的机理,为筛选木质纤维素降解微生物提供一定的指导。同时提出使用合成生物学的策略进行相应微生物的筛选,旨在为筛选高效降解木质纤维素的微生物提供一定的参考。     

酿酒酵母人造纤维小体的研究进展

纤维素乙醇的统合生物加工过程(consolidated bioprocessing,CBP)是将(半)纤维素酶生产、纤维素水解和乙醇发酵过程组合,通过一种微生物完成的生物加工过程。CBP有利于降低生物转化过程的成本,受到研究者的普遍关注。酿酒酵母(Saccharomyces cerevisiae)作为传统的乙醇生产菌株,是极具潜力的CBP底盘细胞。纤维小体是某些厌氧微生物细胞表面由纤维素酶系与支架蛋白组成的大分子复合物,它能高效降解木质纤维,在酿酒酵母表面展示纤维小体已成为构建CBP细胞的研究热点。笔者综述了人造纤维小体在酿酒酵母细胞表面展示组装的研究进展,重点阐述了纤维小体各元件的设计和改造,并针对酿酒酵母分泌途径的改造,提出提高人造纤维小体分泌组装的可能性策略。     

大肠杆菌对木质纤维素水解液抑制物的胁迫耐受性

木质纤维素类生物质是前景广阔的化石原料替代品，其生物炼制可生产生物能源、生物基化学品和生物材料等多种产品，可降低碳排放，有助于实现“双碳”目标，因此受到越来越多的关注。然而，木质纤维素生物炼制需要经过预处理、微生物发酵和产物纯化等多个步骤，其中，预处理过程产生的多种化合物抑制微生物的细胞生长和发酵性能，是制约生物转化效率的瓶颈之一。大肠杆菌是木质纤维素生物炼制常用的宿主，被广泛应用于多种化合物的生产，研究其对木质纤维素水解液中抑制物的耐受性，对于提高木质纤维素生物炼制效率具有重要意义。本文首先介绍了木质纤维素的主要成分和基本结构，对木质纤维素的预处理方法以及预处理后水解液中的主要抑制物种类进行了简单阐述；随后，总结了木质纤维素水解液中几类主要抑制物呋喃类、羧酸类和酚类对大肠杆菌细胞的毒性，以及大肠杆菌对上述抑制物的胁迫响应机制和基于机制的菌株改造靶点；最后，综述了提高大肠杆菌对上述抑制物的胁迫耐受性的菌株改造策略，包括随机突变、实验室适应性进化和组学辅助的理性设计等，为利用代谢工程构建用于木质纤维素生物炼制的高效大肠杆菌菌株提供参考。     

纤维小体在燃料乙醇中的应用

纤维小体在木质纤维素的降解中起着重要作用。它不仅含有降解纤维素所需的各种纤维素酶系,而且组装成具有高效催化活性的多酶复合体形式。介绍了纤维小体基本结构与功能,重点概述了其在生物燃料乙醇中的应用并对纤维小体的研究提出了展望。     

High-Throughput Screening of Multispecies Biofilm Formation and Quantitative PCR-Based Assessment of Individual Species Proportions,Useful for Exploring Interspecific Bacterial Interactions

Multispecies biofilms are predominant in almost all natural environments, where myriads of resident microorganisms interact with each other in both synergistic and antagonistic manners. The interspecies interactions among different bacteria are, despite the ubiquity of these communities, still poorly understood. Here, we report a rapid, reproducible and sensitive approach for quantitative screening of biofilm formation by bacteria when cultivated as mono- and multispecies biofilms, based on the Nunc-TSP lid system and crystal violet staining. The relative proportion of the individual species in a four-species biofilm was assessed using quantitative PCR based on SYBR Green I fluorescence with specific primers. The results indicated strong synergistic interactions in a four-species biofilm model community with a more than 3-fold increase in biofilm formation and demonstrated the strong dominance of two strains, Xanthomonas retroflexus and Paenibacillus amylolyticus. The developed approach can be used as a standard procedure for evaluating interspecies interactions in defined microbial communities. This will be of significant value in the quantitative study of the microbial composition of multispecies biofilms both in natural environments and infectious diseases to increase our understanding of the mechanisms that underlie cooperation, competition and fitness of individual species in mixed-species biofilms.     

Curcumin and wikstroflavone B,a new biflavonoid isolated from Wikstroemia indica,synergistically suppress the proliferation and metastasis of nasopharyngeal carcinoma cells via blocking FAK/STAT3 signaling pathway

BackgroundCurcumin (CUR) is a natural diarylheptanoid with marked anti-tumor activities. Recent investigations demonstrate that CUR combines with some other phytochemicals exerts advantages over its single application manifested as lower toxicity, higher efficacy or more significant reversal of multidrug resistance.PurposeThis study aimed to elucidate a new biflavonoid (wikstroflavone B, WFB) isolated from Wikstroemia indica and to assess the synergistic inhibition of combined CUR and WFB (CUR/WFB) on human nasopharyngeal carcinoma (NPC) cell lines proliferation and metastasis.MethodsWFB was obtained through sequential chromatographic methods including silica gel, Sephadex LH-20 and preparative HPLC. Its structure was determined by HRESIMS, 1D and 2D NMR spectroscopic analysis. The absolute configuration of WFB was assigned through comparison of experimental and calculated optical rotation (OR) values. Changes in cellular viability, migration and invasion were assessed by MTT, colony formation, wound healing and Transwell assays. The nature of synergistic interaction of CUR/WFB was determined through the combination index (CI) method under the median-effect analysis. Expression levels of indicated mRNAs and proteins were measured by qRT-PCR and Western blotting assays, respectively.ResultsWFB was isolated and structural elucidated. Compared with CUR or WFB used alone, CUR/WFB treatment inhibited more effectively on the cell viability, colony formation, cell migration and invasion. Both CI and dose reduction index (DRI) values indicated the significant synergistic effects existed between CUR and WFB. Besides, CUR/WFB showed the marked modulation on the genes involved in cell proliferation (survivin, cyclin D1, p53 and p21) and metastasis (MMP-2, MMP-9 and FAK). CUR/WFB treatment was also found to restrain the phosphorylation of FAK and STAT3 proteins. When pretreatment with a FAK inhibitor, the cell viability and metastasis were significantly attenuated.ConclusionThe results indicate that WFB can synergistically increase the inhibitory effects of CUR on NPC cells proliferation and metastasis, and these findings may afford a rational approach for developing the antitumor medications.     

What Does It Take to Synergistically Combine Sub-Potent Natural Products into Drug-Level Potent Combinations?

There have been renewed interests in natural products as drug discovery sources. In particular, natural product combinations have been extensively studied, clinically tested, and widely used in traditional, folk and alternative medicines. But opinions about their therapeutic efficacies vary from placebo to synergistic effects. The important questions are whether synergistic effects can sufficiently elevate therapeutic potencies to drug levels, and by what mechanisms and at what odds such combinations can be assembled. We studied these questions by analyzing literature-reported cell-based potencies of 190 approved anticancer and antimicrobial drugs, 1378 anticancer and antimicrobial natural products, 99 natural product extracts, 124 synergistic natural product combinations, and 122 molecular interaction profiles of the 19 natural product combinations with collective potency enhanced to drug level or by >10-fold. Most of the evaluated natural products and combinations are sub-potent to drugs. Sub-potent natural products can be assembled into combinations of drug level potency at low probabilities by distinguished multi-target modes modulating primary targets, their regulators and effectors, and intracellular bioavailability of the active natural products.     

Recent advances in natural antifungal flavonoids and their derivatives

The resistance of pathogenic fungi and failure of drug therapy increased dramatically. Numerous studies have reported the individual or synergistic antifungal potency of natural and synthesized flavonoids, especially against drug-resistant fungi. This brief review summarizes the structure and individual or synergistic antifungal activity of natural and synthesized flavonoids (literatures mainly cover the past 10 years 2009–2019), with a special focus on the antifungal spectra, structure–activity relationship and mechanisms of actions. These may contribute to a better understanding of flavonoids as multi-target agents in the treatment of mycoses and provide some ideas on the development of novel flavonoids-based antifungals.     

An optimized biotechnological system for the production of centellosides based on elicitation and bioconversion of Centella asiatica cell cultures

Centella asiatica is a herbaceous plant of Asian traditional medicine. Besides wound healing, this plant is recommended for the treatment or care of various skin conditions such as dry skin, leprosy, varicose ulcers, eczema, and/or psoriasis. Triterpene saponins, known as centellosides, are the main metabolites associated with these beneficial effects. Considering the interest in these high value active compounds, there is a need to develop biosustainable and economically viable processes to produce them. Previous work using C. asiatica plant cell culture technology demonstrated the efficient conversion of amyrin derivatives into centellosides, opening a new way to access these biomolecules. The current study was aimed at increasing the production of centellosides in C. asiatica plant cell cultures. Herein, we report the application of a new elicitor, coronatine, combined with the addition of amyrin‐enriched resins as potential sustainable precursors in the centelloside pathway, for a positive synergistic effect on centelloside production. Our results show that coronatine is a powerful elicitor for increasing centelloside production and that treatments with sustainable natural sources of amyrins enhance centelloside yields. This process can be scaled up to an orbitally shaken CellBag, thereby increasing the capacity of the system for producing biomass and centellosides.     

A reverse chemical ecology approach to explore wood natural durability

The natural durability of wood species, defined as their inherent resistance to wood-destroying agents, is a complex phenomenon depending on many biotic and abiotic factors. Besides the presence of recalcitrant polymers, the presence of compounds with antimicrobial properties is known to be important to explain wood durability. Based on the advancement in our understanding of fungal detoxification systems, a reverse chemical ecology approach was proposed to explore wood natural durability using fungal glutathione transferases. A set of six glutathione transferases from the white-rot Trametes versicolor were used as targets to test wood extracts from seventeen French Guiana neotropical species. Fluorescent thermal shift assays quantified interactions between fungal glutathione transferases and these extracts. From these data, a model combining this approach and wood density significantly predicts the wood natural durability of the species tested previously using long-term soil bed tests. Overall, our findings confirm that detoxification systems could be used to explore the chemical environment encountered by wood-decaying fungi and also wood natural durability.     

SOME EFFECTS OF ANTIBIOTICS IN SURGERY

Although surgical mortality has decreased considerably in the antibiotic era, infectious complications still head the list of causes of death after operation.The choice of an antibiotic singly or in synergistic combinations should be governed by both the pathogenic organism and the organ involved. Allergy, toxicity and development of bacterial resistance are important limiting factors.Besides oxygen, antibiotics are indicated in dealing with localized or generalized anoxia as encountered in vascular impairment of intestinal obstruction or in shock.In acute abdominal conditions meticulous observation is required during the use of antibiotics in order to avoid dangerous delay of operation.     

On adaptation, the assessment of adaptations, and the value of adaptive arguments in phylogenetic reconstruction

Evolutionary adaptation concerns a relative concept and the study of adaptations is directed to structures of individuals. The concept is devoid of any meaning when it is applied to species or populations. Adaptation is not synonymous with fitness or survival but does contribute to both of them. The term adaptation has a dual meaning since it refers both to the process of adaptation and to the state of being adapted. In the process of adaptation the mechanism of natural selection takes a prominent position. But the operation and effectiveness of natural selection are constrained by various limiting factors. Besides that, features may also be the result of nonadaptive evolution and only attain their present adaptive function at a later point in time. Another possibility is that features have at present a function different from the one for which they were initially designed. With respect to the state of being, the study of adaptation attempts to examine whether a particular feature indeed forms an adequate response to selection forces from the environment. Five methods or approaches generally are used to assess the adaptive significance of features, viz. the comparative, correlation, optimization, cladistic, and synthetic approach. Only the last-mentioned approach forms an adequate method since it attempts to establish, by direct analysis, which well-defined selection force exerts its influence on a certain character. The practicing taxonomist is faced with the problem that the data necessary to apply the synthetic method, generally require detailed field studies. Not all evolutionary changes are under the influence of natural selection. The presence of some features may be based on entirely different mechanisms, such as genetic drift, mutational pressure, pleiotropic gene action, allometric growth, or ecophenotypic responses. Various problems inherent to the optimization approach, and several others of practical and theoretical nature, make the morphocline method of the functional and evolutionary morphologists unsuitable as a method for phylogenetic reconstruction.     

Enhanced activity of antifungal drugs using natural phenolics against yeast strains of Candida and Cryptococcus

Aims: Determine whether certain, natural phenolic compounds enhance activity of commercial antifungal drugs against yeast strains of Candida and Cryptococcus neoformans. Methods and Results: Twelve natural phenolics were examined for fungicidal activity against nine reference strains of Candida and one of C. neoformans. Six compounds were selected for synergistic enhancement of antifungal drugs, amphotericin B (AMB), fluconazole (FLU) and itraconazole (ITR). Matrix assays of phenolic and drug combinations conducted against one reference strain, each, of Candida albicans and C. neoformans showed cinnamic and benzoic acids, thymol, and 2,3‐ and 2,5‐dihydroxybenzaldehydes (‐DBA) had synergistic interactions depending upon drug and yeast strain. 2,5‐DBA was synergistic with almost all drug and strain combinations. Thymol was synergistic with all drugs against Ca. albicans and with AMB in C. neoformans. Combinations of benzoic acid or thymol with ITR showed highest synergistic activity. Of 36 combinations of natural product and drug tested, none were antagonistic. Conclusions: Relatively nontoxic natural products can synergistically enhance antifungal drug activity, in vitro. Significance and Impact of the Study: This is a proof‐of‐concept, having clinical implications. Natural chemosensitizing agents could lower dosages needed for effective chemotherapy of invasive mycoses. Further studies against clinical yeast strains and use of animal models are warranted.     

Systems biology, connectivity and the future of medicine

The concept of systems-based strategies in medicine is emerging, with systems pathology guiding an understanding of the multidimensional aspects of disease system fingerprints and systems pharmacology providing insight into dynamic system responses upon (multiple) drug perturbations. Knowledge of the changes of system characteristics during disease progression creates a framework for the design of novel combinatorial treatment strategies. Such a systems-based, combinatorial-therapies approach readdresses the value of the synergistic actions of components of treatments based on natural products and highlights new methodology to study multidimensional intervention via reversed-pharmacology.