Engineering lysine‐rich caleosins as carrier proteins to render biotin as a hapten on artificial oil bodies for antibody production

It has been demonstrated that caleosin alone is sufficient to stabilize artificial oil bodies. A series of recombinant caleosins, mutated with 3, 5, 8, 11, 13, 15, and 17 extra Lys residues and over‐expressed in Escherichia coli, were used as carrier proteins to render biotin as a hapten on the surface of artificial oil bodies for antibody production. Biotinylation levels of the recombinant caleosins were step‐wisely elevated as the number of extra Lys residues increased, and the biotinylated Lys residues were identified by mass spectrometric analysis. Polyclonal antibodies against biotin were successfully generated in rats injected with artificial oil bodies constituted with each of the biotinylated caleosins. Moreover, those generated via the biotinylated caleosins with eight or more extra Lys residues no longer recognized caleosin. It appears that engineered Lys‐rich caleosins are suitable carrier proteins for the production of antibodies against small molecules. © 2011 American Institute of Chemical Engineers Biotechnol. Prog., 2011     

Acetylation of SUMO1 Alters Interactions with the SIMs of PML and Daxx in a Protein-Specific Manner

1. Download : Download high-res image (112KB)
2. Download : Download full-size image

     

Adaptable P body physical states differentially regulate bicoid mRNA storage during early Drosophila development

1. Download : Download high-res image (265KB)
2. Download : Download full-size image

     

The morphology of multivesicular bodies in soybean protoplasts and their role in endocytosis

Summary Multivesicular bodies (MVBs) in soybean protoplasts are distinct organelles (generally 250–500 nm in diameter) consisting of a limiting membrane and a number of smaller internal vesicles (generally 40–100 nm in diameter). MVBs of soybean protoplasts are morphologically similar to MVBs of animal cell systems. They can have tubular protuberances which extend from the main body of the organelle and a lamellar plaque on the cytoplasmic surface of their limiting membrane. In addition, the internal vesicles can be labeled by a zinc iodide-osmium tetroxide postfixation and may form via invagination of the limiting membrane.The MVBs of soybean protoplasts are a major compartment in the endocytotic pathway. They accumulate, over time, exogenously applied cationized ferritin and may deliver it to the major lysosomal or lytic compartment of the plant cell, namely, the vacuoles.     

Ketone Body Utilization for Lipid Synthesis in the Murine Sciatic Nerve: Alterations in the Dysmyelinating Trembler Mutant

This work demonstrates that in vitro sciatic nerves of normal and trembler adult mice can use ketone bodies (beta-hydroxybutyrate and acetoacetate) and butyrate for lipid synthesis. In normal sciatic nerves, beta-hydroxybutyrate is incorporated in total lipids to a larger extent than acetoacetate (141% and 33%, respectively, of acetate incorporation), whereas for trembler sciatic nerves, these percentages are only 69% and 27%. Incorporation of ketone bodies is greater into sterols than into other lipids. Lipid metabolism of ketone bodies in trembler nerves is altered and could reflect a process similar to Wallerian degeneration: a dramatic decrease of sterol and free fatty acid synthesis and an increased synthesis of triglycerides. Moreover, differences seen in precursor incorporation into lipids between normal and trembler sciatic nerves suggest that their lipid metabolism is not the same.     

Woronin bodies in arthrinium genus

Woronin bodies are present near all septal pores and in conidia of Arthrinium strains and may regulate cytoplasmic flow in both injured and actively growing healthy colonies. They vary in size and frequency, the central one plugs the septal pores in actively developing colonies and in mature conidia. The septa are thinner in the Woronin-body region.     

Reserve substances inPaxillus involutus sclerotia

Summary The ectomycorrhizal fungus,Paxillus involutus, produces sclerotia in culture. These can be induced to form on agar medium by exposing mycelium grown at 25°C to various temperatures between6°C and 15°C. Sclerotia formed at 10°C and above were large and covered with drops of exudate, while those formed at 6°C or 8°C were very small and did not produce an exudate. Mature sclerotia were bounded by a compact rind and contained abundant storage reserves. Histochemistry of the larger sclerotia showed large quantities of protein stored as protein bodies in the cytoplasm, lipid present as small droplets, glycogen granules stored in the cytoplasm and polyphosphate present as small granules in the cytoplasm and in the protein bodies. Energy dispersive X-ray microanalysis confirmed the presence of phosphate in the granules and was used to map its distribution throughout the sclerotium. The smaller sclerotia induced at 8°C and below on the same medium had the same basic structure and composition, but lacked the complex phenolic cell network found in large sclerotia, and had abundant extracellular polysaccharides. The rind was not well developed and these small sclerotia are interpreted to have been arrested at an early stage of development.     

Ultrastructural visualization of exocytosis in the pig pineal gland

Summary The pinealocytes of the pig contain conspicuous dense bodies, the nature and role of which are not yet fully elucidated. The aim of this study was to demonstrate whether or not these structures are involved in the secretion process. The tannic acid-Ringer incubation (TARI)-method, which allows a clear-cut ultrastructural study of secretory discharge by exocytosis, has been used. The results indicate that pig pinealocytes release the content of the dense bodies with an amorphous inner structure into the extracellular space via exocytosis and that this secretion is quantitatively important. The secreted material is proteinaceous in nature; this indicates that polypeptides are released by the pineal.     

Purification of ribulose 1,5-bisphosphate carboxylase/oxygenase and of carboxysomes from Thiobacillus thyasiris the putative symbiont of Thyasira flexuosa (Montagu)

The bacterial symbionts of many marine invertebrates contain ribulose 1,5-bisphosphate (RuBP) carboxylase but apparently no carboxysomes, polyhedral bodies containing RuBP carboxylase. In the few cases where polyhedral bodies have been observed they have not been characterised enzymatically. Polyhedral bodies, 50–90 nm in diameter, were observed in thin cell sections of Thiobacillus thyasiris the putative symbiont of Thyasira flexuosa and RuBP carboxylase activity was detected in both soluble and particulate fractions after centrifugation of cell-free extracts. RuBP carboxylase purified 90-fold from the soluble fraction was of high molecular weight and consisted of large and small subunits, with molecular weights of 53,110 and 11,100 respectively. Particulate RuBP carboxylase activity was associated with polyhedral bodies 50–100 nm in diameter, as revealed by density gradient centrifugation and electron microscopy. Therefore, the polyhedral bodies were inferred to be carboxysomes. Native electrophoresis of isolated carboxysomes demonstrated a major band which comigrated with the purified RuBP carboxylase and three minor bands of lower molecular weight. Sodium dodecyl-sulphate (SDS) gel electrophoresis of SDS-dissociated carboxysomes demonstrated nine major polypeptides two of which were the large and small subunits of RuBP carboxylase. The RuBP carboxylase subunits represented 21% of the total carboxysomal protein. The most abundant polypeptide had a molecular weight of 40,500. Knowledge of carboxysome composition is necessary to provide an understanding of carboxysome function.Abbreviations FPLC fast performance liquid chromatography - IB isolation buffer - PAGE polyacrylamide gel electrophoresis - RuBP carboxylase - ribulose 1,5-bisphosphate carboxylase/oxygenase - SDS sodium dodecyl-sulphate     

一例硬皮病人自发抗核仁抗体的免疫荧光及其与银染的比较研究

从硬皮病人血清中筛选出一例含有自发抗核仁抗体的血清,利用这个血清对核仁抗原的性质及其在细胞中随分裂周期不同产生的分布变化做了初步研究,并把结果与核仁嗜银蛋白做了比较。间接免疫荧光染色及细胞化学分析表明,这种核仁抗原的性质是蛋白质,其分布与嗜银蛋白相似,在间期,抗原呈颗粒状簇集在核仁中,而在分裂中期,抗原颗粒与染色体NORs部位接合,但有证据指出,这种抗原蛋白与核仁嗜银蛋白有所不同,同时还发现,经长时间秋水仙素处理诱导产生微核化的多核细胞中尽管微核的数目远多于细胞中NORs的数目,免疫荧光染色和银染都显示出每个微核中类核仁小体的存在。这说明(1)类核仁小体也是由核仁物质构成;(2)某些类核仁小体的产生可能与NORs无关。对这个现象的意义进行了讨论。