淀粉接枝丙烯酰胺与丙烯酸的实验研究

在氮气保护下，以N，N’-亚甲基双丙烯酰胺为交联剂，过硫酸铵和亚硫酸钠作为引发剂，淀粉与丙烯酰胺、丙烯酸接枝共聚。经干燥后制备吸去离子水迭700倍的吸水性树脂。探讨了反应时间、交联剂用量、引发剂用量、单体中和度、反应温度、淀粉用量、干燥温度对吸水倍率的影响。     

添加辣椒碱类化合物合成抗菌高吸水树脂

以过硫酸铵、亚硫酸氢钠作为引发剂，N，N’-亚甲基双丙烯酰胺作为交联剂，将木薯淀粉与丙烯酸、辣椒碱在水溶液中接枝共聚，合成了具有抗菌性的高吸水性树脂。文章主要考察了添加辣椒碱形式、反应温度、反应时间、引发剂用量、丙烯酸中和度、交联剂用量对合成产物吸水性能的影响。     

蔗渣纤维丙烯酸高吸水树脂的制备

以蔗渣纤维为原料,丙烯酸为接枝单体,N,N-亚甲基双丙烯酰胺为交联剂,K2S2O8为引发剂,通过溶液聚合的方法合成了蔗渣纤维（sugarcanebagasse,SCB）与聚丙烯酸（polyacrylicacid,PAA）的接枝共聚高吸水树脂（scB—g—PAA）。研究了丙烯酸用量、引发剂用量、交联剂用量以及中和度对吸水树脂吸水倍率的影响,结果表明,该接枝共聚吸水树脂的最佳合成条件是以丙烯酸用量为标准,蔗渣纤维、K2S2O8、交联剂用量分别是10％、2％、0．1％,中和度为80％,共聚物吸水倍率最高为765．7g·g-1。     

玉米秆制取吸水树脂的研究

以玉米秆为原料制取羧甲基纤维素钠,与丙烯酸接枝共聚制备吸水树脂.讨论了单体配比、丙烯酸中和度、交联剂用量及引发剂用量等因素对吸水性能的影响.实验发现,在丙烯酸单体与玉米秆质量比为8∶1、丙烯酸中和度为70%、交联剂N,N-亚甲基双丙烯酰胺为丙烯酸单体质量的0.24%、引发剂过硫酸钾为丙烯酸单体质量的3.5%、反应温度为60℃的条件下,所得树脂吸水倍率最高可达68 g/g.     

接枝共聚法制备高吸水性树脂

采用反相悬浮聚合法,以过硫酸钾为引发剂,N,N-亚甲基双丙烯酰胺为交联剂,以淀粉为主要原料,与丙烯酰胺、丙烯酸接枝共聚制备了高吸收性树脂。讨论了搅拌速率、分散剂的种类、引发剂用量、交联剂用量、丙烯酸的中和度、环己烷与水的比例对接枝共聚物吸水性能的影响。制备得到的树脂吸水能力可达600g／g,吸0．9％的生理盐水达70g／g。     

羧甲基纤维素、淀粉双母体超强吸水树脂的合成及性能研究

采用自由基聚合法，以过硫酸钾为引发剂，N，N-亚甲基双丙烯酰胺为交联剂，羧甲基纤维素、玉米淀粉与丙烯酸钠进行自由基接枝共聚合成吸水剂。并详细考察了中和度、原料配比、引发剂和交联剂的用量对接枝共聚物性能的影响。通过实验得到合成该吸水保水剂的适宜条件。实验结果表明。该接枝共聚物是一种较好的超强吸水材料。     

淀粉接枝共聚合成吸水树脂工艺研究

以淀粉为原料,丙烯酸和丙烯酰胺为共聚单体,过硫酸铵为引发剂,N,N‘-亚甲基双丙烯酰胺为交联剂,采用接枝共聚法制备吸水树脂。研究了引发剂用量、交联剂量、单体与淀粉质量比以及丙烯酸和丙烯酰胺质量比对产品的吸水率的影响。实验结果表明：当引发剂占干物料质量的0．6％,交联剂质量占单体质量的0．8‰,单体与淀粉质量比为4,丙烯酸与丙烯酰胺质量比为1．33,产品的吸水率可超过550。     

麦秸秆纤维素与丙烯酸接枝共聚制备耐盐性吸水树脂的研究

应用小麦秸秆与丙烯酸接枝共聚制备了耐盐性吸水树脂,进行了结构表征,研究了单体配比、丙烯酸中和度、引发剂和交联剂用量以及反应温度对吸盐水倍率的影响。研究发现,接枝共聚的适宜条件为:丙烯酸单体与麦秸秆质量比为8∶1,丙烯酸中和度为70%,引发剂过硫酸钾-硫代硫酸钠的用量为单体的3.5%,交联剂N,N-亚甲基双丙烯酰胺用量为单体质量的0.24%,反应温度为70℃。在此条件下制备的树脂吸盐水倍率最高,吸盐水(CNaC l=0.9%)可达68 g/g,可应用于医疗卫生等方面。     

羧甲基纤维素、玉米淀粉双母体含氮高吸水树脂的合成及性能测定

采用自由基聚合法,以过硫酸铵为引发剂,N,N'-亚甲基双丙烯酰胺为交联剂,羧甲基纤维素及玉米淀粉与丙烯酸进行自由基接枝共聚合成吸水剂.并详细考察了原料配比、中和度、引发剂和交联剂的用量对接枝共聚物性能的影响,通过实验得到该吸水保水剂的适宜条件.     

机械活化花生壳-丙烯酸/丙烯酰胺水凝胶的制备及性能研究

利用机械活化后的花生壳粉末为基质,丙烯酸/丙烯酰胺为复合单体,过硫酸钾作引发剂,N,N一亚甲基双丙烯酰胺作交联剂,在水溶液中接枝聚合制备复合水凝胶。讨论了丙烯酸中和度、机械活化时间、单体配比、引发剂用量、交联剂用量、反应温度和反应时间对水凝胶的吸水倍率的影响。实验结果表明,在较佳的工艺条件下水凝胶对去离子水和生理盐水吸水倍率分别为148.37 g/g和19.78 g/g。     

The influence of the region between residues 220 and 344 and beyond in Phrixotrix railroad worm luciferases green and red bioluminescence

To find the regions having a major influence on the bioluminescence spectra of railroad worm luciferases, we constructed new chimeric luciferases switching the fragments from residues 1-219 and from 220-545 between Phrixotrix viviani (PxvGR; lambda(max) = 548 nm) green light-emitting luciferase and Phrixothrix hirtus (PxhRE; lambda(max) = 623 nm) red light-emitting luciferases. The emission spectrum (lambda(max) = 571 nm) and K(M) for luciferin in the chimera PxRE220GR (1-219, PxhRE; 220-545, PxvGR) suggested that the region above residue 220 of PxvGR had a major effect on the active site. However, switching the sequence between the residues 220-344 from PxvGR luciferase into PxhRE (PxREGRRE) luciferase resulted in red light emission (lambda(max) = 603 nm), indicating that the region 220-344 by itself does not determine the emission spectrum. Furthermore, the sequence before residue 220 of the green-emitting luciferase is incompatible for light emission with the sequence above residue 220 of PxhRE. These results suggest that the fragments before and after residue 220, which correspond to distinct subdomains, may fold differently in the green- and red-emitting luciferases, affecting the active site conformation.     

Conservation and covariance in PH domain sequences: physicochemical profile and information theoretical analysis of XLA-causing mutations in the Btk PH domain

Mutations that cause X-linked agammaglobulinemia (XLA) appear throughout the Bruton tyrosine kinase (Btk) sequence, including the pleckstrin homology (PH) domain. To analyze the basis of this disease with respect to protein structure, we studied the relationships between PH domain sequences and structures by comparing sequence-based profiles of physicochemical properties and solvent accessibility profiles. The diversity of the distribution of amino acids was measured by calculating entropies for sequences containing mutations at different positions in multiple sequence alignments. Mutual information was calculated to quantify positional covariation. Eight conserved extrema were apparent in all profiles. The majority of the XLA disease-causing mutations in the Btk PH domain were found at positions having significant mutual information, indicating that there are covariant constraints for both structure and function. Together with additional structural analyses, all the XLA mutations that were analyzed could be explained at the molecular level. The method developed here is applicable to the design of mutations for protein engineering.     

Improved activity and thermostability of Candida antarctica lipase B by DNA family shuffling

DNA family shuffling was used to create chimeric lipase B proteins with improved activity toward the hydrolysis of diethyl 3-(3',4'-dichlorophenyl)glutarate (DDG). Three homologous lipases from Candida antarctica ATCC 32657, Hyphozyma sp. CBS 648.91 and Crytococcus tsukubaensis ATCC 24555 were cloned and shuffled to generate a diverse gene library. A high-throughput screening assay was developed and used successfully to identify chimeric lipase B proteins having a 20-fold higher activity toward DDG than lipase B from C.antarctica ATCC 32657 and a 13-fold higher activity than the most active parent derived from C.tsukubaensis ATCC 24555. In addition, the stability characteristics of several highly active chimeric proteins were also improved as a result of family shuffling. For example, the half-life at 45 degrees C and melting point (T(m)) of one chimera exceeded those of lipase B from C.antarctica ATCC 32657 by 11-fold and 6.4 degrees C, respectively, which closely approached the stability characteristics of the most thermostable parent derived from Hyphozyma sp. CBS 648.91.     

An efficient method for creation and functional analysis of libraries of hybrid type I polyketide synthases

Bacterial type I polyketide synthases (PKSs) generate a structurally diverse group of natural products with a wide range of biological activities. Hybrid type I PKSs in which domains of one multifunctional polypeptide are replaced with components from heterologous systems have generated significant interest over the past decade. Almost invariably only one or several specific hybrids are made at a time and tested for functionality. This approach is slow, dependent upon a fortuitous choice of specific fusions points, and often leads to inactive or minimally active hybrid systems. We describe herein a method for generating and screening a library of hybrid pikAI complementation plasmids (encoding the loading domain and the first two extension domains of pikromycin PKS) able to restore pikromycin in a BB138 Streptomyces venezuelae pikAI-deletion mutant. In the first step the plasmid sequence encoding the loading domain AT(0)-ACP(0) was replaced by a counter selectable marker, sacB. DNA family shuffling was then used to generate a diverse library of chimeric AT(0)-ACP(0) fragments, which were used to replace sacB by lambda-Red-mediated in vivo recombination in an Escherichia coli host. This method resulted in the rapid and efficient generation of a large number of hybrid pikAI complementation plasmids, which were used to transform S.venezuelae BB138. A bioassay of over 4000 of these transformants successfully revealed three different PikAI hybrids which were able to lead to pikromycin production. The study suggests that most of the hybrids are not detectably functional, and underscores the need to generate and screen large and diverse libraries in which different fusion points are tried. The methodologies applied in this study address this need and can be used for directed evolution of any component of the PikPKS, and potentially other type I PKS systems.     

浅谈现代塑料模具的设计与制造

随着现代塑料制品的形状越来越复杂，塑料模具的设计也越来越复杂。以载重汽车仪表板为例，介绍了鞠料模具设计的内容和过程。介绍了CAD／CAE／CAM、先进设备、手工加工、检测手段、反向工程、快速成型制造在模具制造中的应用。分析了现代塑料模具制造的发展方向和前景。     

A novel emulsion mixture for in vitro compartmentalization of transcription and translation in the rabbit reticulocyte system

Emulsion formulations used for in vitro compartmentalization (IVC) methods were found to be incompatible with protein expression in the rabbit reticulocyte (RRL) system, causing rapid discoloration and translation shutdown. Here we identify possible causes and describe a novel water-in-oil emulsion which abolished discoloration and allowed high-level in-emulsion expression of active luciferase and human telomerase using the RRL. This novel emulsion greatly expands the range of potential protein targets for IVC.     

Fusion of the antiferritin antibody VL domain to barnase results in enhanced solubility and altered pH stability

Chimeric immunotoxins that combine antigen recognition domains of antibodies and cytotoxic RNases have attracted much attention in recent years as potential targeted agents for cancer immunotherapy. In an attempt to obtain a structurally minimized immunofusion for folding/stability studies, we constructed the chimeric protein VL-barnase. The chimera comprises a small cytotoxic enzyme barnase, ribonuclease from Bacillus amyloliquefaciens, fused to the C-terminus of the light chain variable domain (VL) of the anti-human ferritin monoclonal antibody F11. While the individual VL domain was expressed in Escherichia coli as insoluble protein packed into inclusion bodies, its fusion to barnase resulted in a significant ( approximately 70%) fraction of soluble protein, with only a minor insoluble fraction ( approximately 30%) packed into inclusion bodies. The in vivo solubilizing effect of barnase was also observed in vitro and suggests a chaperone-like role that barnase exerted with regard to the N-terminal VL domain. Cytoplasmic VL-barnase was analyzed for structural and functional properties. The dimeric state of the chimeric protein was demonstrated by size-exclusion chromatography, thus indicating that fusion to barnase did not abrogate the intrinsic dimerization propensity of the VL domain. Ferritin-binding affinity and specificity in terms of constants of association with isoferritins were identical for the isolated VL domain and its barnase fusion, and RNase activity remained unchanged after the fusion. Intrinsic fluorescence spectra showed a fully compact tertiary structure of the fusion protein. However, significantly altered pH stability of the fusion protein versus individual VL and barnase was shown by the pH-induced changes in both intrinsic fluorescence and binding of ANS. Together, the results indicate that VL-barnase retained the antigen-binding affinity, specificity and RNase activity pertinent to the two individual constituents, and that their fusion into a single-chain chimeric protein resulted in an altered tertiary fold and pH stability.     

Transglutaminase-mediated N- and C-terminal fluorescein labeling of a protein can support the native activity of the modified protein

Fluorescein and its analogs are among the best fluorophores to label proteins and the labeling generally involves chemical modification of a translated protein. Using this methodology, labeling at a specific position remains difficult. It is known that the guinea pig liver transglutaminase (TGase)-catalyzed enzymatic modification method can allow terminal-specific fluorophore labeling of a protein by monodansylcadaverine. However, native activity of the fluorescent protein has not been investigated so far, nor has direct comparison between the chemical modification and the TGase-catalyzed modification been attempted. Therefore, we compared the possibility of fluorescein labeling via chemical labeling and via TGase-catalyzed modification. The latter method was found to be very practical and overcame some of the problems associated with the specificity of the former; fluorescein was covalently attached only to the N- or C-terminal site of glutathione S-transferase when the reaction was catalyzed by TGase and the resulting labeled protein completely retained its native activity. The TGase-mediated labeling occurred not only at room temperature but also at 4 degrees C to the same extent, which is more desirable for preventing the inactivation of proteins.     

阴离子交换树脂分离双波长分光光度法测定铅

在 p H7.5～ 9.0的缓冲介质中 ,铅与 5 - Br- PADAP形成稳定的红色三元胶束络合物 ,以 440 nm为参比波长 ,5 70 nm为测定波长 ,可用双波长法测定微量铅 ,表观摩尔吸光系数为ε=1 .0× 1 0 5L· mol- 1· cm- 1,铅的浓度在 0～ 1 .6mg· L- 1范围内符合比尔定律。本法用于矿样中微量铅的测定 ,结果较满意