Article 1：植物细胞壁表面的纤维素和木质素共定位限制了微生物对杨树生物质的水解
Title：Cellulose and lignin colocalization at the plant cell wall surface limits microbial hydrolysis of Populus biomass.
DOI: 10.1039/c7gc00346c.
Abstract: Biorefining of plant feedstocks into fuels and specialty chemicals, using biological conversion, requires the solubilization of lignocellulosics into simpler oligomeric compounds. However, non-pretreated woody biomass has shown high resistance to hydrolysis by cellulolytic microbes or purified cellulases. We investigate the limited solubilization of Populus deltoides by the cellulolytic thermophile Clostridium thermocellum in the absence of solute inhibitors. Compared to control samples, fermented poplar revealed that the hydrolysis of carbohydrates in secondary cell walls ceased prematurely as the presence of lignin increased at the surface. In quantitative fluorescence colocalization analysis by confocal laser scanning microscopy, the Manders’coefficient of the fractional overlap between lignin and cellulose signals increased from an average of 0.67 to a near-maximum of 0.92 in fermented tissue. Chemical imaging by time-of-flight secondary ion mass spectrometry revealed a 49% decline in surface cellulose and a compensatory 30% and 11% increase in surface S- and G-lignin, respectively. Although 72% of the initial glucan was still present in the lignocellulose matrix of this feedstock, subsequent treatments with cell-free purified cellulases did not significantly restore hydrolysis. This confirmed that biomass surfaces had become non-productive for the C. thermocellum hydrolytic exoproteome. This study provides direct evidence for an explicit definition of feedstock recalcitrance, whereby depletion of surface carbohydrate increases lignin exposure which leads to inhibition of enzyme activity, while the bulk residual biomass retains significant undigested carbohydrate content. The analysis presented here establishes a novel method for the quantitation of lignocellulose recalcitrance.
研究背景：木质纤维素中的纤维素分为结晶区和无定形区。热纤梭菌 (Clostridium thermocellum) 是一种快速生长的厌氧嗜热纤维素分解细菌，能够在难降解的木质纤维素材料上生长,分泌多种细胞结核酶和游离糖苷水解酶，能将结晶纤维素有效水解至接近完全。
难点：在没有对原料进行预处理的情况下，很难解聚植物细胞壁中的复杂木质纤维素。虽然最近的研究集中在木质素对抑制真菌纤维素酶的影响上，但是木质素和热纤梭菌之间的相互作用还需要进一步研究，特别是在表面木质素方面。
研究内容：文章中假设植物组织表面的共定位和木质素和纤维素的相对比例的变化导致水解活性过早停止。 为了验证这一点，设置对照组（有/无C. thermocellum 参与水解），使用共聚焦激光扫描显微镜 (CLSM) 的定量荧光分析比较了 60 µm 厚的水解杨树切片与非水解组的对照，并通过Time-of-Flight二次离子质谱（TOF-SIMS）分析了表面化学变化。
Result analysis：
Article 2：使用荧光蛋白标记的碳水化合物结合模块评估干燥对纤维素可及性和酶水解的影响
Title：The use of fluorescent protein-tagged carbohydrate-binding modules to evaluate the influence of drying on cellulose accessibility and enzymatic hydrolysis.
DOI: 10.1039/d0ra05333c.
Abstract: The influence of drying on cellulose accessibility and enzymatic hydrolysis was assessed. Dissolving pulp was differentially dried by freeze-, air- and oven-drying at 50℃and subsequently hydrolyzed using the commercial CTec 3 cellulase preparation. It was apparent that drying reduced the ease of enzymatic hydrolysis of all of the substrates with a pronounced reduction (48%) exhibited by the oven-dried pulp. To assess if the ease of hydrolysis was due to enzyme accessibility to the substrate, microscopy (SEM), FTIR spectroscopy, water retention value (WRV), fiber aspect ratio analysis, Simons' stain and the selective binding of Fluorescent Protein-tagged Carbohydrate Binding Modules (FP-CBMs): CBM3a (crystalline cellulose) and CBM17 (amorphous cellulose) in combination with confocal laser scanning microscopy (CLSM) were used. The combined methods indicated that, if the gross characteristics of the substrate limited enzyme accessibility, the cellulases, as represented by the FP-CBMs, could not in turn access the finer structural components of the cellulosic substrates.
研究背景：研究表明，木质纤维素酶解分解成糖的限速步骤是纤维素酶进入纤维素组分的能力（酶的可及性），影响因素包括粒度、聚合度、木质素和半纤维素分布等。
难点：确定纤维素可及性已被证明很难用在宏观和微观尺度上用纤维素底物在非晶化阶段期间的酶介导动态情况来评估。
实验方法：CBMs是碳水化合物活性酶的非活性多糖识别区，如糖苷水解酶。通常，CBMs被分为三种类型：a、b和c，基于它们的三维结构和功能相似性，a型识别结晶纤维素，b型和c型分别识别内部聚糖链(内切型)和末端(外切型)聚糖。
研究内容：研究表明干燥会导致木质纤维素发生实质性改变。这篇文章中，CBMs被用于在结构水平上的量化和定位可及性， 提供纤维素底物的易接近的无定形和结晶区域的区别。用于评估干燥对纤维素酶可及性的影响。
Result analysis：
Article 3：一种基于电致变色普鲁士蓝膜和双极电极系统中固定化葡萄糖氧化酶的生物催化的可复位和可重编程键盘锁
Title：A resettable and reprogrammable keypad lock based on electrochromic Prussian blue films and biocatalysis of immobilized glucose oxidase in a bipolar electrode system
DOI: 10.1016/j.bios.2017.07.054
文章原理图：
实验结论：
总结：
这种键盘锁定系统结合了酶催化和双极电化学，显示出一些独特的优势，如良好的可重编程性，易于复位和肉眼可视读数。基于葡萄糖氧化酶酶催化系统，可以在机理创新性和应用方面进一步研究。
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