Alpha-factor-leader-directed secretion of recombinant human-insulin-like growth factor I from Saccharomyces cerevisiae. Precursor formation and processing in the yeast secretory pathway

A synthetic gene coding for human-insulin-like growth factor I (IGFI) was fused to the leader sequence of yeast prepro-alpha-factor and expressed in Saccharomyces cerevisiae under the control of a glyceraldehyde-3-phosphate dehydrogenase promoter fragment. Recombinant IGFI was found inside yeast cells and secreted into the medium. The secreted IGFI migrated on SDS gels with the same electrophoretic mobility as authentic IGFI, i.e. at about 7.5 kDa. HPLC analysis of secreted IGFI revealed the presence of the correctly folded, genuine molecule as well as an isomeric byproduct of equal molecular mass but with two of the three disulfide bonds interchanged. Inside exponentially growing cells the 7.5-kDa IGFI was also found, along with up to four additional IGFI-related polypeptides of higher molecular mass. By endoglycosidase F treatment the three polypeptides between 19-26 kDa were converted to a single peptide of 17 kDa. Since this peptide also reacted with an anti-alpha-factor antibody, it represents most likely the unglycosylated alpha-factor--IGFI fusion precursor. Pulse-chase experiments established the precursor nature of the intracellular higher-molecular-mass IGFI species. Conversion of the primary translation product to the differently glycosylated IGFI precursor proteins and into the mature form occurred very rapidly, within 2 min. Rapid maturation was, however, not followed by an equally rapid secretion of the mature form into the medium: only after 30-40 min did IGFI appear outside the cells. We therefore postulate the presence of an as yet undefined Golgi or post-Golgi bottleneck representing a major obstacle in secretion of recombinant IGFI from S. cerevisiae cells.     

Isolation and characterisation of a crystalline antithiamine factor from mustard seed, Brassica juncea

The isolation and characteristics of a crystalline antithiamine factor from mustard seed, Brassica juncea is described.This oil bearing seed seems to contain at least two antithiamine factors, one of these is the methyl ester of 3,5-dimethoxy 4-hydroxy cinnamic acid (methyl sinapate). The chemical nature of the other is not yet known because it could not be isolated in pure form as yet.     

A new name for Limnophyes montanus Chaudhuri,Sinharay & Das Gupta (Diptera: Chironomidae) from India

Limnophyes nigripes is proposed as a replacement name for Limnophyes montanus Chaudhuri, Sinharay & Das Gupta, nee Limnophyes montanus Goetghebuer.     

Deformable models for segmentation of CLSM tissue images and its application in FISH signal analysis.

In this paper we present an application of deformable models for the segmentation of volumetric tissue images. The three-dimensional images are obtained using confocal microscope. The segmented images have been used for the quantitative analysis of the Fluorescence In Situ Hybridization (FISH) signals. An ellipsoidal surface initialized around the cell of interest acts as a deformable model. The deformable model surface voxels are subjected to various internal and external forces derived from underlying image features as well as externally imposed constraints. The deformable model converges to the optimum cell shape when the vector sum of all the forces acting on the model is zero. The result of segmentation is used to confirm the cell membership of the FISH signals and to reject all the signals that lie outside the cell nuclei. Three-dimensional region isolation and labeling technique is used to label and count the FISH signals per cell nucleus. A simple study on the effect of different segmentation methods over a quantitative analysis of FISH signals is also presented.     

Production and excretion of nitrate by human newborn infants: neonates are not little adults.

Endothelium-derived relaxing factor, identified as nitric oxide or its adducts, is metabolized to nitrate and excreted in the urine. Since blood pressures are lower in newborn infants compared to adults, we hypothesized that newborn infants would have increased excretion of nitrate on the day of birth. Neonatal urine was collected before 24 h of age when exogenous intake of nitrate was low. Two different analytical methods showed that nitrate accounted for >99% of nitrogen oxides in urine of healthy neonates and adults. The absolute micromolar concentration of nitrate in urine from infants was significantly below that of adults. When nitrate content was standardized for the reduced renal function in the newborn infant (creatinine content) and body mass (kilogram weight), the concentration of nitrate in neonatal urine was significantly higher than that of adults. Nitrate concentrations in the urine of prematurely born infants were twice that of nitrate measured in urine from term infants. These findings suggested that nitric oxide is produced in larger intravascular quantities in newborn infants versus adults. Thus, we postulated that nitric oxide released from a nitrosothiol would be metabolized to nitrate more readily by neonatal erythrocytes compared to red blood cells obtained from adults. Neonatal erythrocytes, suspended at concentrations of 8, 12, or 16 g per deciliter of hemoglobin, produced 1.7- to 2.1-fold more nitrate than equivalent hemoglobin concentrations of adult erythrocytes that were each incubated with S-nitroso-N-acetylpenicillamine (100 microM) over a 2-h period. Taken together, the studies of urinary nitrate in newborn infants and the ability of neonatal erythrocytes to generate nitrate are consistent with a robust production of nitric oxide immediately after birth.     

Novel non-glycerol-based cytofectins with lactic acid-derived head groups.

We report herein the design, synthesis, and transfection biology of a novel series of non-glycerol-based cationic lipids with lactic acid-derived head groups The synthetic procedure adopted herein for preparing 1-hydroxy-prop-2-yl head-group-based monocationic transfection lipids 1-7 is fairly straightforward and potentially applicable in designing other cationic lipids with lactic acid-derived head groups. A striking anchor-length dependency was observed in NIH3T3 cells in the sense that except lipid 4, all the other lipids were essentially transfection-inefficient. Ethidium bromide assay for the lipid:DNA interactions is consistent with the general observation that significant lipid:DNA interactions do not guarantee on improved transfection efficiency cationic lipid mediated gene delivery. Given its remarkable transfection properties and low cellular toxicity, lipid 4 is likely to find future use in the area of liposomal gene delivery.     

Nanoantenna-Enhanced Radiative and Anisotropic Decay Rates in Monolayer-Quantum Dots

Plasmonics - Nanoantenna-enhanced ultrafast emission from colloidal quantum dots as quantum emitters is required for fast quantum communications. On-chip integration of such devices requires a...