Immobilization of yeast cells containing beta-galactosidase

Cultural conditions optimum for beta-galactosidase production by Saccharomyces anamensis are pH 4.5, temperature 26 +/- 2 degrees C, and 30 h of incubation period. Addition of lactose at 24 h fermentation greatly increase the level of enzyme. Optimum pHl, temperature, pH stability, and thermostability of yeast beta-galactosidase are negligibly affected by immobilization. The K(m) values of enzyme in the native and immobilized cells are 102mM and 148mM, respectively. Glucose noncompetitively inhibits the enzyme activity. Addition of substances such as dithioerythritol, glutathione, and bovine serum albumin to the native cell during assay procedure and immobilized cell prior to immobilization have stimulatory effects on enzyme activity. Metal ions like Ca(2+), Mg(2+) enhance the beta-galactosidase activity for both intact and bound cells. Immobilized cells retain 68.6% of the beta-galactosidase activity of intact cells and there is no significant loss of activity on storage at 4 degrees C for 28 days.     

Changes in lipid profiles of brain of albino rats following envenomation

Effect of high doses of cobra venom (150 micrograms/120 +/- 20 g body weight) and viper venom (300 micrograms/120 +/- 20 g body weight) on total lipid, triglyceride, phospholipid, cholesterol, high density lipoprotein cholesterol (HDL-C) and low density lipoprotein cholesterol (LDL-C) of brain of albino rats was studied. Total lipid (TL) triglyceride (TG) and phospholipid (PL) are decreased in both viper and cobra venom treated groups while cholesterol (C), and LDL-C are increased in both the groups in relation to controlled ones. HDL-C content was almost unaltered. Decrease in triglyceride and phospholipid may be due to effect of lipases and phospholipases whereas increased cholesterol and LDL-C may be attributed to lysis of cell membrane.     

Inhibition of D-glucose uptake by isatin in rat intestine: effect of harmaline and various sulfhydryl reagents

The effect of isatin (indole-2,3-dione) on D-glucose uptake has been studied in rat intestine. Isatin at 6 mM concentration significantly inhibited both the sugar uptake and transmural (mucosal to serosal side) transport in the intestine. The suppression of glucose uptake by isatin was irreversible. Similar to the action of various SH-group-reacting agents, isatin inhibited the sugar uptake, presumably by binding to membrane sulfhydryl groups through a covalent linkage. Isatin-induced reduction in glucose uptake was unaffected by pH (between 5.5 and 8.4) and by DTT addition to incubation medium. Inhibition of sugar uptake by isatin and harmaline was additive in nature; this suggested that these compounds interact at different sites on the microvillus membrane surface.     

De‐ubiquitinases on the move: an emerging field in plant biology

A balance between the synthesis and degradation of active proteins governs diverse cellular processes in plants, spanning from cell‐cycle progression and circadian rhythm to the outcome of several hormone signalling pathways. Ubiquitin‐mediated post‐translational modification determines the degradative fate of the target proteins, thereby altering the output of cellular processes. An equally important, and perhaps under‐appreciated, aspect of this pathway is the antagonistic process of de‐ubiquitination. De‐ubiquitinases (DUBs), a group of processing enzymes, play an important role in maintaining cellular ubiquitin homeostasis by hydrolyzing ubiquitin poly‐proteins and free poly‐ubiquitin chains into mono‐ubiquitin. Further, DUBs rescue the cellular proteins from 26S proteasome‐mediated degradation to their active form by cleaving the poly‐ubiquitin chain from the target protein. Any perturbation in DUB activity is likely to affect proteostasis and downstream cellular processes. This review illustrates recent findings on the biological significance and mechanisms of action of the DUBs in Arabidopsis thaliana, with an emphasis on ubiquitin‐specific proteases (UBPs), the largest family among the DUBs. We focus on the putative roles of various protein–protein interaction interfaces in DUBs and their generalized function in ubiquitin recycling, along with their pre‐eminent role in plant development.     

Glutamatergic Neurotransmission from Melanopsin Retinal Ganglion Cells Is Required for Neonatal Photoaversion but Not Adult Pupillary Light Reflex

Melanopsin-expressing retinal ganglion cells (mRGCs) in the eye play an important role in many light-activated non-image-forming functions including neonatal photoaversion and the adult pupillary light reflex (PLR). MRGCs rely on glutamate and possibly PACAP (pituitary adenylate cyclase-activating polypeptide) to relay visual signals to the brain. However, the role of these neurotransmitters for individual non-image-forming responses remains poorly understood. To clarify the role of glutamatergic signaling from mRGCs in neonatal aversion to light and in adult PLR, we conditionally deleted vesicular glutamate transporter (VGLUT2) selectively from mRGCs in mice. We found that deletion of VGLUT2 in mRGCs abolished negative phototaxis and light-induced distress vocalizations in neonatal mice, underscoring a necessary role for glutamatergic signaling. In adult mice, loss of VGLUT2 in mRGCs resulted in a slow and an incomplete PLR. We conclude that glutamatergic neurotransmission from mRGCs is required for neonatal photoaversion but is complemented by another non-glutamatergic signaling mechanism for the pupillary light reflex in adult mice. We speculate that this complementary signaling might be due to PACAP neurotransmission from mRGCs.     

BUD13 Promotes a Type I Interferon Response by Countering Intron Retention in Irf7

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Characterization of a galactose specific adhesin of enteroaggregative Escherichia coli

A fimbrial adhesin was identified from an enteroaggregative Escherichia coli strain. The adhesin was purified to 740-fold by sequential chromatography on an affinity matrix and gel filtration column in the FPLC system. The homogeneity of the purified protein was established by analytical isoelectrofocussing (pI 7.25). The native adhesin appeared as a high-molecular-weight aggregative protein as revealed by gel filtration chromatography on Superose 12HR10/30 column. However, in sodium dodecyl sulfate-polyacrylamide gel electrophoresis the molecular weight of the adhesin was found to be 18 kDa and this was further confirmed by gel filtration chromatography on Superose 6HR 10/30 column presence of 6 M guanidine hydrochloride. The N-terminal 15-amino-acid sequence of the adhesin did not show homology with any of the previously reported fimbrial adhesins. The purified adhesin showed adhesion to human erythrocytes in the presence of Ca(2+) (5 mM). The optimum temperature and pH for the hemadhesion activity was found to be 25 degrees C and 6.5, respectively. The inhibition study clearly suggested that the binding site of the adhesin could recognize galactose as the specific sugar. The fluorescence of 4-methylumbelliferyl-alpha-D-galactopyranoside was quenched on binding to the adhesin and maximum reversal of fluorescence quenching was observed by competitive substitution titration with raffinose. The adhesin was found to contain one binding site per monomer for its specific sugar residue. The association constant and the free energy of binding were obtained as 3.98 x 10(5) M(-1) and -31.97 kJ/mol, respectively. The adherence of the bacteria to HEp-2 monolayer was inhibited in presence of galactose and this was further supported by a significant reduction in the bacterial adherence to the HEp-2 cells, pretreated with beta-D-galactosidase.