过敏原肌质钙结合蛋白在毕赤酵母中的表达、优化及免疫反应性研究

目的 探究天然肌质钙结合蛋白(sarcoplasmic calcium binding protein, SCP)的可替代物,为蟹类过敏原的检测提供基础材料,本研究首次利用毕赤酵母(Pichia pastoris, P. pastoris)高效表达表达三疣梭子蟹(Portunus trituberculatus)重要过敏原SCP,并检验其免疫反应性。方法 根据毕赤酵母的密码子偏好性优化SCP基因并构建重组质粒。将其热激转化至P. pastoris GS115菌株后经遗传霉素(Geneticin, G418)筛选获得阳性高拷贝子。最后通过甲醇诱导表达重组SCP并结合免疫印记(Western blotting, WB)和间接酶联免疫吸附实验(enzyme-linked immunosorbent assay, ELISA)验证其免疫反应性。结果 SCP在P. pastoris GS115中实现了可溶性高效表达,其表观分子量约为28 kDa。在摇瓶水平下,最佳诱导条件为pH为6.0、每24 h添加1.0%(v/v)甲醇,于28℃发酵144 h,在此条件下,纯度为91.6%的SCP产量可达15 mg/L。WB和间接ELISA结果表明,重组SCP具有IgG结合能力。结论 毕赤酵母表达系统可以得到纯度较高且免疫反应性良好的重组SCP。本研究为SCP的理化研究及产业化应用奠定了基础,并有望促进特异性甲壳类过敏原检测的发展。     

A Constraint-Based Approach for Deriving 3-D Structures of Cyclic Polypeptides

This paper presents an hybrid algorithm for deriving 3-D structures of cyclic polypeptides. The algorithm combines constraint-based techniques with the most widely used methods for non-cyclic polypeptides. The empirical results demonstrate that the proposed hybrid algorithm outperforms traditional methods especially with respect to running times.     

用同步辐射X荧光分析血浆蛋白质的微量元素

用同步辐射X荧光(SRXRF)分析技术与凝胶色谱生化分离相结合，分析了用抗肿瘤药物顺铂处理过的和未处理过的小鼠血浆的大分子蛋白质。对以轻元素为基体的小量样品，用X荧光谱中的康普顿散射强度作为质量的因子，获得了在大于22ku分子量的蛋白质中Pt、S、Ca、Fe、Ni、Cu、Zn、Se、Br和Sr等元素含量在不同样品问的相对变化。由此推测，经顺铂处理后，Pt元素可与血浆中的大分子蛋白结合，含CM和S的大分子蛋白含量明显升高，而含Zn的减少。     

大豆/涤混纺纱混纺比及其不匀率的测试与计算方法

为了研究大豆蛋白纤维这种新型的纺织原料混纺比的测试方法,利用数理统计原理,采用密度梯度法对大豆/涤纶(65/35)混纺纱混纺比进行了测试和计算,该方法不仅可以得到混纺比均值,而且可以得到混纺比标准差,粗纱和细纱混纺比均值的控制范围分别为(63.64%,65.32%)和(64.16%,66.17%),粗纱和细纱混纺比标准差的控制限分别为2.89%和3.56%,同期细纱用溶解法实验结果为64.5%,2种方法误差为1.1%。     

河南省市售鱼虾蟹类及其制品营养成分调查分析

目的 分析河南省市售及加工鱼虾蟹类食品的营养成分,完善河南省特色食物成分数据库。方法 2017年12月—2020年12月对饭店和超市内售卖的鱼蟹类食品采样,分析其中的水分、灰分、蛋白质、脂肪、碳水化合物、总氮、糖类、多种维生素及人体必需的矿物质等营养成分。结果 水分、灰分含量水平基本一致;除油炸食品外均未检出糖类;蟹类能量含量整体偏高,炸鱼能量、蛋白质、总氮和脂肪整体高于鲜鱼;鱼虾蟹类脂肪酸的总量不高,不饱和脂肪酸比例大,二十碳五烯酸（EPA）和二十二碳六烯酸（DHA）占主要成分,DHA含量较高的依次是鱿鱼0.209 g/100 g、龙虾0.181 g/100 g和巴沙鱼0.153 g/100 g,EPA依次是蟹黄1.07 g/100 g、小黄鱼0.38 g/100 g和鲫鱼0.297 g/100 g;鱼类全部检出氨基酸,虾蟹类90%检出,蟹类中蟹黄氨基酸含量较高。结论 鱼虾蟹类制品中,调味料及食用油对食物成分有影响;鲜品中不饱和脂肪酸的含量与总脂肪酸含量正相关,鱼类和蟹类单不饱和脂肪酸高,虾类与推荐比例一致。     

Structural Insights into Iron Ions Accumulation in Dps Nanocage

Dps (DNA-binding protein from starved cells) is well known for the structural protection of bacterial DNA by the formation of highly ordered intracellular assemblies under stress conditions. Moreover, this ferritin-like protein can perform fast oxidation of ferrous ions and subsequently accumulate clusters of ferric ions in its nanocages, thus providing the bacterium with physical and chemical protection. Here, cryo-electron microscopy was used to study the accumulation of iron ions in the nanocage of a Dps protein from Escherichia coli. We demonstrate that Fe²⁺ concentration in the solution and incubation time have an insignificant effect on the volume and the morphology of iron minerals formed in Dps nanocages. However, an increase in the Fe²⁺ level leads to an increase in the proportion of larger clusters and the clusters themselves are composed of discrete ~1–1.5 nm subunits.     

Secreted HSP90α-LRP1 Signaling Promotes Tumor Metastasis and Chemoresistance in Pancreatic Cancer

The extracellular heat shock protein 90α (eHSP90α) has been reported to promote cancer cell motility. However, whether pancreatic cancer (PC) cells expressed membrane-bound or secreted HSP90α, as well as its underlying mechanism for PC progression, were still unclear. Our study demonstrated that the amounts of secreted HSP90α proteins were discrepant in multiple PC cells. In addition, highly invasive Capan-2 cells have a higher level of secreted HSP90α compared with those of less invasive PL45 cells. The conditioned medium of Capan-2 cells or recombinant HSP90α treatment stimulated the migration and invasion of PC cells, which could be prevented with a neutralizing anti-HSP90α antibody. Furthermore, secreted HSP90α promoted elements of epithelial–mesenchymal transition in PL45 cells, including increases in vimentin and Snail expressions, decreases in E-cadherin expression, and changes in cell shape towards a mesenchymal phenotype, but these phenomena were reversed by the anti-HSP90α antibody in Capan-2 cells. In addition, high levels of low-density lipoprotein receptor-related protein 1 (LRP1) were associated with worsened patient survival in pancreatic adenocarcinoma. We demonstrated LRP1 as a receptor of eHSP90α for its stimulatory role in metastasis, by activating the AKT pathway. In addition, silencing LRP1 enhanced the chemosensitivity to gemcitabine and doxorubicin in Capan-2 cells. Therefore, our study indicated that blocking secreted HSP90α underlies an aspect of metastasis and chemoresistance in PC.     

Lysozyme Amyloid Fibril Structural Variability Dependence on Initial Protein Folding State

Amyloid fibril formation is associated with several amyloidoses, including neurodegenerative Alzheimer’s or Parkinson’s diseases. The process of such fibrillar structure formation is still not fully understood, with new mechanistic insights appearing on a regular basis. This, in turn, has limited the development of potential anti-amyloid compounds, with only a handful of effective cures or treatment modalities available. One of the multiple amyloid aggregation factors that requires further examination is the ability of proteins to form multiple, structurally distinct aggregates, based on the environmental conditions. In this work, we examine how the initial folding state affects the fibrilization of lysozyme—an amyloidogenic protein, often used in protein aggregation studies. We show that there is a correlation between the initial state of the protein and the aggregate formation lag time, rate of elongation, resulting aggregate structural variability and dye-binding properties, as well as formation lag time and rate of elongation.     

The Potential Role of SP-G as Surface Tension Regulator in Tear Film: From Molecular Simulations to Experimental Observations

The ocular surface is in constant interaction with the environment and with numerous pathogens. Therefore, complex mechanisms such as a stable tear film and local immune defense mechanisms are required to protect the eye. This study describes the detection, characterization, and putative role of surfactant protein G (SP-G/SFTA2) with respect to wound healing and surface activity. Bioinformatic, biochemical, and immunological methods were combined to elucidate the role of SP-G in tear film. The results show the presence of SP-G in ocular surface tissues and tear film (TF). Increased expression of SP-G was demonstrated in TF of patients with dry eye disease (DED). Addition of recombinant SP-G in combination with lipids led to an accelerated wound healing of human corneal cells as well as to a reduction of TF surface tension. Molecular modeling of TF suggest that SP-G may regulate tear film surface tension and improve its stability through specific interactions with lipids components of the tear film. In conclusion, SP-G is an ocular surface protein with putative wound healing properties that can also reduce the surface tension of the tear film.