碳酸酐酶和甲酸脱氢酶的稳定性研究进展

化石燃料的大量燃烧使温室气体CO₂的排放量不断增加,对环境造成恶劣影响,将CO₂捕集并转化为高附加值化学品是实现节能减排和变废为宝的一种双赢策略。酶催化CO₂捕集和转化具有高效、高选择性、反应条件温和、环境友好等优点。碳酸酐酶（CA）可大大加速CO₂水合反应,而甲酸脱氢酶（FDH）可催化CO₂还原为甲酸,二者协同可增强CO₂还原动力学。但酶促反应的工业化应用过程中,酶所处环境的温度、酸碱度以及其他离子的种类和浓度等因素均可能导致酶失活,因此,酶的稳定性研究至关重要。本文从热稳定性、酸碱稳定性和离子稳定性的角度,综述了CA和FDH的稳定性研究进展。改善酶稳定性的手段包括使用极端微生物、酶分子设计与改造、固定化等,重点讨论了固定化对酶稳定性的提升效果,为未来的工业化应用提供参考。     

Dendrimer as nanocarrier for drug delivery

Dendrimers are novel three dimensional, hyperbranched globular nanopolymeric architectures. Attractive features like nanoscopic size, narrow polydispersity index, excellent control over molecular structure, availability of multiple functional groups at the periphery and cavities in the interior distinguish them amongst the available polymers. Applications of dendrimers in a large variety of fields have been explored. Drug delivery scientists are especially enthusiastic about possible utility of dendrimers as drug delivery tool. Terminal functionalities provide a platform for conjugation of the drug and targeting moieties. In addition, these peripheral functional groups can be employed to tailor-make the properties of dendrimers, enhancing their versatility. The present review highlights the contribution of dendrimers in the field of nanotechnology with intent to aid the researchers in exploring dendrimers in the field of drug delivery.     

Teriflunomide Preserves Neuronal Activity and Protects Mitochondria in Brain Slices Exposed to Oxidative Stress

Teriflunomide (TFN) limits relapses in relapsing–remitting multiple sclerosis (RRMS) by reducing lymphocytic proliferation through the inhibition of the mitochondrial enzyme dihydroorotate dehydrogenase (DHODH) and the subsequent modulation of de novo pyrimidine synthesis. Alterations of mitochondrial function as a consequence of oxidative stress have been reported during neuroinflammation. Previously, we showed that TFN prevents alterations of mitochondrial motility caused by oxidative stress in peripheral axons. Here, we aimed to validate TFN effects on mitochondria and neuronal activity in hippocampal brain slices, in which cellular distribution and synaptic circuits are largely preserved. TFN effects on metabolism and neuronal activity were investigated by assessing oxygen partial pressure and local field potential in acute slices. Additionally, we imaged mitochondria in brain slices from the transgenic Thy1-CFP/COX8A)S2Lich/J (mitoCFP) mice using two-photon microscopy. Although TFN could not prevent oxidative stress-related depletion of ATP, it preserved oxygen consumption and neuronal activity in CNS tissue during oxidative stress. Furthermore, TFN prevented mitochondrial shortening and fragmentation of puncta-shaped and network mitochondria during oxidative stress. Regarding motility, TFN accentuated the decrease in mitochondrial displacement and increase in speed observed during oxidative stress. Importantly, these effects were not associated with neuronal viability and did not lead to axonal damage. In conclusion, during conditions of oxidative stress, TFN preserves the functionality of neurons and prevents morphological and motility alterations of mitochondria.     

Modeling Alcohol Dehydrogenase Catalysis in Deep Eutectic Solvent/Water Mixtures

The use of oxidoreductases (EC1) in non-conventional reaction media has been increasingly explored. In particular, deep eutectic solvents (DESs) have emerged as a novel class of solvents. Herein, an in-depth study of bioreduction with an alcohol dehydrogenase (ADH) in the DES glyceline is presented. The activity and stability of ADH in mixtures of glyceline/water with varying water contents were measured. Furthermore, the thermodynamic water activity and viscosity of mixtures of glyceline/water have been determined. For a better understanding of the observations, molecular dynamics simulations were performed to quantify the molecular flexibility, hydration layer, and intraprotein hydrogen bonds of ADH. The behavior of the enzyme in DESs follows the classic dependence of water activity (a_W) in non-conventional media. At low a_W values (<0.2), ADH does not show any activity; at higher a_W values, the activity was still lower than that in pure water due to the high viscosities of the DES. These findings could be further explained by increased enzyme flexibility with increasing water content.     

甲酸脱氢酶用于辅酶NADH再生的研究进展

NADH依赖型氧化还原酶广泛应用于精细化学品和手性化合物的生物合成,其中辅酶NADH作为还原当量起着关键的作用,NADH的再生关系到生物氧化还原过程能否进行. 甲酸脱氢酶在辅酶NADH再生中的应用是目前代谢工程领域研究的热点之一. 本工作回顾了甲酸脱氢酶的来源和氨基酸序列、酶的理化性质和催化机理等方面的研究进展,从化学稳定性、热稳定性和成本等方面阐述了甲酸脱氢酶在辅酶再生系统中的应用,讨论了甲酸脱氢酶用于辅酶再生的代谢工程平台的发展趋势,并对研究发展方向提出了一些设想.     

生物转化法生产木糖醇的细胞增殖培养基优化

以木糖为底物利用酵母细胞转化生产木糖醇,研究细胞增殖培养基中不同种类的碳源、氮源和微量元素及其添加量对酵母细胞生长和木糖转化率的影响。结果表明,当碳源为木糖和葡萄糖添加量分别为1%时,木糖醇浓度为49.6g/L;当无机氮源为蛋白胨且浓度为2%时,木糖转化时间为60h,木糖醇浓度达到58g/L;微量元素为磷酸二氢钾浓度为0.1%时,木糖的转化时间为48h,此时木糖醇浓度达到59g/L。     

Green Tea Formulations with Vitamin C and Xylitol on Enhanced Intestinal Transport of Green Tea Catechins

The effect of green tea formulated with vitamin C and xylitol on intestinal cell transport of gallated and nongallated catechin was studied. The transport of catechins from both apical to basolateral and basolateral to apical directions was measured. The effect of vitamin C (4, 10, 20 ppm), xylitol (11, 27.5, 55 ppm), and combinations of both on the intestinal transport rate of catechins was examined. The efflux value (Pb→a/Pa→b) of (–)‐epigallocatechin (EGC), (–)‐epigallocatechin gallate (EGCG), (–)‐epicatechin (EC), and (–)‐epicatechin gallate (ECG) was 0.26, 0.22, 1.22, and 0.17, respectively, indicating that EC appeared to be less absorbed compared with other catechins. The addition of xylitol (11, 27.5, 55 ppm) and vitamin C (4, 10, 20 ppm) and in combination enhanced transport rate of nongallated catechins such as EC and EGC. For EC, vitamin C was revealed to be the most effective on intestinal transport, implying the inhibition of the efflux transport mechanism of EC. Intestinal transport of gallated catechins significantly increased from catechins formulated with vitamin C and xylitol in a dose‐dependent manner compared to the catechin‐only formulation. Results provide a potential strategy to enhance the delivery and bioavailability of catechins in humans by modulating green tea formulation with vitamin C and xylitol.     

三种糖醇对高粱淀粉糊化特性和凝胶结构的影响

本文通过快速黏度分析仪(RVA)、物性测试仪(TPA)、差示扫描热量仪(DSC)、扫描电子显微镜(SEM),研究不同浓度的赤藓糖醇、木糖醇、麦芽糖醇(淀粉与糖醇比例为1:1、1:2、1:5,m/m),对高粱淀粉性质和高粱淀粉凝胶结构的影响。与原高粱淀粉相比,随着三种糖醇添加量的增加,相同温度下高粱淀粉的溶胀度下降,尤其是在75℃、85℃和95℃。RVA结果表明,随着糖醇添加量增加,高粱淀粉的峰值黏度和衰减值下降,然而其糊化温度增加。糖醇使高粱淀粉凝胶的硬度升高,而且随着糖醇添加量增加而增加。DSC结果表明,添加三种糖醇后,高粱淀粉的起始糊化温度,峰值糊化温度,终止糊化温度,焓变均增加,而且随着糖醇添加量增加而升高。从SEM可知,添加糖醇后的高粱淀粉凝胶的微观结构变得更紧密。     

重组L-乳酸脱氢酶在大肠杆菌中的表达、纯化及活性研究

乳酸脱氢酶是生物法转化苯丙酮酸为苯乳酸的一种有效的酶。实验克隆到一种新型L-乳酸脱氢酶基因ldhL,来源于Lactobacillus plantarumSK-2(植物乳杆菌SK-2),GenBank接受号为FJ392647。以pET-22b(+)为载体质粒,E.coliBL21(DE3)为宿主细胞,构建了基因重组菌,IPTG可诱导目的蛋白的过量表达;经亲和层析纯化的重组蛋白样品进行SDS-PAGE电泳分析,约在37ku处出现显著的特征蛋白条带;重组LDH的比酶活为0.06U/mg。     

利用TAIL-PCR克隆Gluconobacter suboxydans葡萄糖脱氢酶基因及其生物信息学分析

以Gluconobacter suboxydans J菌株基因组DNA为模板,基于吡咯喹啉醌附着位点的保守区域设计引物,通过交错式热不对称PCR获得编码葡萄糖脱氢酶（glucose dehydrogenase,GDH）的全长基因（gdh）,并对其序列进行生物信息学分析。结果表明：gdh基因全长2 268 bp,编码755个氨基酸,其蛋白序列与Gluconobacter属的GDH具有较高的同源性。GDH的分子质量约为81.72 ku,pI值约为5.14;GDH蛋白的二级结构由18.41%的α-螺旋、16.16%的延伸和65.43%的无规则卷曲3种结构模块组成;GDH N末端的AA 1～140区域有5个跨膜结构域。