聚乙二醇修饰的单克隆抗体的某些生物活性

 不久前我们报道了PEG-1900修饰的McAb的某些理化性质,本文主要报道经PEG-1900修饰的McAb在某些生物活性方面的变化。与天然的McAb相比,修饰的McAb有以下的变化:(1)抗原性与免疫原性下降;(2)抗体活性下降;(3)在体内存活的时间延长;(4)对温度及pH的抵抗力增强。修饰的酶与天然的酶相比,酶活性有所下降,但下降的程度要比修饰的McAb小得多。     

Structure and function of the microtubular cytoskeleton during endosperm development in wheat: an immunofluorescence study

Summary The three-dimensional structure of the microtubular cytoskeleton of developing wheat endosperm was investigated immunocytochemically. Semi-thin sections were prepared from polyethylene glycol embedded ovaries. At the free-nuclear stage the endosperm cytoplasm with regularly distributed nuclei surrounded a large central vacuole and exhibited an extensive network of fluorescent labelled microtubular assemblies radiating from each nucleus. As was found in other coenocytes, this particular and nuclear-dependent cytoskeletal configuration functions in the arrangement of nuclei and in the stabilization of the nuclear positions. At the beginning of cellularization of the endosperm the formation of vacuoles altered the radiating networks. It is likely that the radiating microtubular arrays function in the formation of phragmoplasts, independent of nuclear divisions. The formation of anticlinal cell walls, giving rise to openended cell cylinders, coincides with the occurrence of phragmoplast microtubular arrays which were demonstrated during the period of cell wall elongation. The microtubular system radiating from the nuclei in these cell cylinders anchored the nuclei in stage- and locus-specific positions. During the development of aleurone and inner endosperm cells, cell morphogenesis was related to earlier demonstrated types of microtubular configurations in the cortical cytoplasm. This suggests that a general mechanism is involved.Abbreviations A alveolus - AL aleurone layer - CE central endosperm - CV central vacuole - DAP days after pollination - END endosperm - FITC fluorescein isothiocyanate - GAR-FITC goat anti-rabbit antibodies conjugated with FITC - I integument - IE PC inner epidermis pericarp - II inner integument - N nucleus - NC nucellus cells - NE nucellar epidermis - NUC nucellus - OI outer integument - PBS phosphate buffered saline - PC pericarp - PEG polyethylene glycol - V vacuole     

Germination of trinucleated pollen: formulation of a new medium for Capsella bursa-pastoris

Summary In Brewbaker and Kwack's medium (BK) only 16% of the pollen grains germinated, and these produced pollen tubes having a maximum length of 25 m. With a solution based on Monnier's medium 47% germination and 160-mlong pollen tubes were observed. Calcium was shown to be essential for germination; the optimal concentration was 880 mg/l calcium chloride. The optimal concentrations of magnesium sulphate and boric acid were 360 and 50 mg/l, respectively. Germination at pH 4.0 but also pH 8.0 and the presence of vitamins B1 and B6 (1 mg/l each) were stimulatory. Polyethylene glycol (PEG) was superior to sucrose as an osmoticum and germination and tube length were significantly improved using PEG 4000 at a concentration of 120 g/l (0.03 M). Equimolar concentrations of PEG 400 and PEG 600 gave inferior results. Combining PEG with sucrose in the medium did not improve germination or increase tube length.     

The dominant bacteria shifted from the order “ Lactobacillales ” to Bacillales and Actinomycetales during a start-up period of large-scale, completely-mixed composting reactor using plastic bottle flakes as bulking agent

Bacterial community succession in the start-up of a large-scale, completely-mixed composting reactor was analyzed by 16S rRNA gene (16S rDNA) clone analysis and denaturing gradient gel electrophoresis (DGGE) combined with measurements of temperature, pH, moisture contents, and decomposing rate. DGGE analysis and physicochemical parameters showed that bacterial community succession occurred in four phases; (1) at the start of operation and pH decreasing period (day 0–3), (2) pH decreased and increased period (day 4–11), (3) middle term, moisture content decreasing and maximum temperature increased period (day 12–16) and (4) latter term, temperature decreasing period (day 17–24). Lactobacillus spp. and Bacillus coagulans were detected from the initial phase and middle term, respectively. 16S rDNA clone analysis showed that the dominant bacteria shifted from the order “Lactobacillales” to Bacillales and Actinomycetales. The order “Lactobacillales” was unique which may be caused by using the plastic bottle flakes (polyethylene terephthalate, PET) as bulking agent.     

聚乙二醇化重组蛋白药物的质量控制

重组蛋白经聚乙二醇(PEG)化修饰在优化药物代谢动力学和药效学性质的同时,使药物的结构和质量属性变得更为复杂,修饰后的重组蛋白在结构、理化性质和生物学活性等方面与未修饰的重组蛋白相比有较大差异,对其质量控制的研究必须结合品种自身独特的质量属性。现从蛋白质药物质量控制的角度,对PEG化蛋白质药物的重要质量属性的质控难点和相应检测方法进行综述,以期对工艺开发和生产上的实际工作有所帮助。     

Significant differences in the activities of alpha-amylases in the absence and presence of polyethylene glycol assayed on eight starches solubilized by two methods

Starch is a reserve chemical source of the energy of the sun found in plants as a water-insoluble granule that differs in their chemical and physical properties, depending on the source. The granules can be solubilized by heating in water or by treatment with various reagents, such as 1M NaOH. alpha-Amylases are widely distributed enzymes that initiate the hydrolysis of starch into low molecular weight maltodextrins. We recently found that the activities of a single alpha-amylase on two different starches were significantly different. We then determined the activities of Bacillus amyloliquefaciens and porcine pancreas alpha-amylases, using eight different starches, solubilized by two methods: autoclaving at 121 degrees C and 1M NaOH at 20 degrees C. There were significant differences in the activities of both of the amylases on all eight of the starches. Previously, it had been found that polyethylene glycol (PEG) stabilized and activated the activities of both enzymes, using a soluble amylose as the substrate. Addition of PEG to the enzymes greatly increased the activities on the eight starches, but the activities still differed significantly. The different activities with the starches were hypothesized as differences in the amounts of secondary and tertiary structures that are partially retained when the different starches are solubilized; the activities on addition of PEG is hypothesized as the formation of highly active species from a series of less active forms.     

Effect of conformity and contact stress on wear in fixed-bearing total knee prostheses

Ultra high molecular weight polyethylene (PE) remains the primary bearing surface of choice in total knee replacements (TKR). Wear is controlled by levels of cross-shear motion and contact stress. The aim of this study was to compare the wear of fixed-bearing total knee replacements with curved and flat inserts and to test the hypothesis that the flat inserts which give higher contact stresses and smaller contact areas would lead to lower levels of surface wear. A low-conforming, high contact stress knee with a low-medium level of cross shear resulted in significantly lower wear rates in comparison to a standard cruciate sacrificing fixed-bearing knee. The low wear solution found in the knee simulator was supported by fundamental studies of wear as a function of pressure and cross shear in the pin on plate system. Current designs of fixed-bearing knees do not offer this low wear solution due to their medium cross shear, moderate conformity and medium contact stress.     

Studies on the activity and stability of immobilized horseradish peroxidase on poly(ethylene terephthalate) grafted acrylamide fiber

Having been activated with glutaraldehyde, modified poly(ethylene terephthalate) grafted acrylamide fiber was used for the immobilization of horseradish peroxidase (HRP). Both the free HRP and the immobilized HRP were characterized by determining the activity profile as a function of pH, temperature, thermal stability, effect of organic solvent and storage stability. The optimum pH values of the enzyme activity were found as 8 and 7 for the free HRP and the immobilized HRP respectively. The temperature profile of the free HRP and the immobilized HRP revealed a similar behaviour, although the immobilized HRP exhibited higher relative activity in the range from 50 to 60 °C. The immobilized HRP showed higher storage stability than the free HRP.     

Combined effects of polymers and KL4 peptide on surface activity of pulmonary surfactant lipids

Addition of ionic and nonionic polymers can improve the function of therapeutic surfactants in vitro and in vivo, especially under conditions that tend to inhibit surfactant activity. Since surfactant proteins also act to reduce surfactant inhibition, we studied the relative effects of a synthetic peptide (that mimics some of the properties of a surfactant protein), polymers, and their combination on function of surfactant phospholipid activity in vitro. We evaluated surface activity after adding polymers—polyethylene glycol or hyaluronan—to a lipid mixture with or without the synthetic peptide, sinapultide (KL₄). Using a pulsating bubble surfactometer, we measured peptide/polymer effects separately or combined at two peptide concentrations. Phospholipid mixtures, with or without KL₄ or polymers, all demonstrated good surface activity. With serum present as an inhibiting agent, adding either concentration of KL₄ reduced inhibition. Mixtures containing the higher concentration of KL₄ required higher concentrations of serum for inhibition to occur. Adding either polymer to mixtures with KL₄ further decreased susceptibility to inhibition (required higher serum concentrations). In the presence of serum, high molecular weight hyaluronan with KL₄ at 0.4 mg/ml improved surface activity to a greater degree than 0.8 mg/ml KL₄ without polymer. If the beneficial effects of adding polymer to KL₄-lipid mixtures are also borne out in the treatment of experimental lung injury, these peptide-polymer surfactant combinations may eventually prove useful in the treatment of some forms of acute lung injury in humans.     

Exploration of the use of an acylsulfonamide safety-catch linker for the polymer-supported synthesis of hyaluronic acid oligosaccharides

The synthesis of hyaluronic acid oligosaccharides on polyethylene glycol (PEG) using an acylsulfonamide linker has been explored. Hyaluronic acid is a challenging synthetic target that usually involves the condensation of highly disarmed glucuronic acid building blocks. Amine-ended PEG monomethyl ether was efficiently functionalized with a hydroxyl-terminated acylsulfonamide linker. Suitably protected d-glucosamine (GlcN) and d-glucuronic acid (GlcA) monosaccharide building blocks were coupled to the polymer acceptor using the trichloroacetimidate glycosylation method. The sulfonamide safety-catch linker enables simultaneous cleavage of the monosaccharide from the polymer and orthogonal functionalization for further (bio)-conjugation of the sugar sample. Subsequent glycosylation of PEG-bound glycosyl acceptor to generate hyaluronic acid oligosaccharide chain failed. Model glycosylation experiments in solution and on soluble support using the same unreactive acceptors and donors allows for the synthesis of an orthogonally protected hyaluronic acid disaccharide and suggest that the encountered difficulties could be attributed to the presence of the N-acylsulfonamide.