Effect of carbon sources on the synthesis of pectinase by Aspergilli

The synthesis of pectinase is investigated using six species of Aspergillus, with five media differing either in their carbon sources or level of carbon source(s). Five of the six species used, synthesized appreciable amounts of pectinase in the media containing sugars. Pectinase synthesis was highest for A. niger, NCIM 548, with all the sugar containing media. A. foetidus, NCIM 510, was the only one among the organisms studied, that responded well to the medium containing pectin in the absence of additional sugars supplied in the medium.     

Expression of plasmid-encoded aad in Clostridium acetobutylicum M5 restores vigorous butanol production.

Mutant M5 of Clostridium acetobutylicum ATCC 824, which produces neither butanol nor acetone and is deficient in butyraldehyde dehydrogenase (BYDH), acetoacetate decarboxylase, and acetoacetyl-coenzyme A:acetate/butyrate:coenzyme A-transferase activities, was transformed with plasmid pCAAD, which carries the gene aad (R. V. Nair, G. N. Bennett, and E. T. Papoutsakis, J. Bacteriol, 176:871-885, 1994). In batch fermentation studies, aad expression restored butanol formation (84 mM) in mutant M5 without any acetone formation or any significant increase in ethanol production. The corresponding protein (AAD) appeared as a ca. 96-kDa band in a denaturing protein gel. Expression of AAD in M5 resulted in restoration of BYDH activity and small increases in the activities of acetaldehyde dehydrogenase, butanol dehydrogenase, and ethanol dehydrogenase. These findings suggest that BYDH activity in C. acetobutylicum ATCC 824 resides largely in AAD, and that AAD's primary role is in the formation of butanol rather than of ethanol.     

Characterization and Plasmid Profile of Xanthomonas campestris pv. Glycines from Maharashtra, India

Xanthomonas campestris pv. glycines , (Xcg), the causative agent of the bacterial pustule disease of soybean was isolated and characterized. On susceptible soybean the pathogenic isolates displayed characteristic chlorotic lesions around the site of infection within 48 h of inoculation. The pathogenic isolates were found to contain two cryptic plasmids. A smaller plasmid of 1.5 kb and a larger one of size about 25 kb. SDS-PAGE profile of the soluble proteins of the pathogenic isolatess, howed a different pattern compared to that of the non-pathogenic isolates.     

Inhibition of Immune Complex-Induced Inflammation by A small Molecular Weight Selectin Antagonist

The anti-inflammatory effect of a small molecular weight antagonist of P- and E-selectin-dependent cell adhesion was examined. The glycolipid sulphatide was shown to block the adherence of thrombin-activated rat platelets to HL-60 cells. This interaction is known to be dependent on P-selectin. The rat dermal reverse passive Arthus reaction was used to assess the effect of sulphatide on a neutrophil dependent inflammatory response. Sulphatide dosedependently blocked both the vascular permeability increase and cell infiltration after intraperitoneal administration. These results show that a small molecular weight compound which blocks P- and E-selectin dependent adhesion in vitro can effectively block the inflammation due to immune complex deposition. A compound with this type of profile may have therapeutic potential in the treatment of immune complex mediated diseases.     

Cloning of ODAP Degrading Gene and Its Expression as Fusion Protein in Escherichia coli

A gene responsible for the degradation of ß-N-Oxalyl diaminopropionic acid (ODAP) was fused to the maIE gene, which codes for maltose binding protein, by cloning into an expression vector pMAL c2. The gene has been expressed as fusion protein of mol wt approximately 62 kD. It has been purified by affinity chromatography. The fusion protein has been cleaved by an endoprotease factor Xa and the presence of maltose binding protein and the product of the cloned gene confirmed. SDS-PAGE has shown that the product of the ODAP degrading gene is a single polypeptide of mol wt of about 20.7 kD.     

Epidemic isolates of Vibrio cholerae 0139 express antigenically distinct types of colonization pili

Abstract Vibrio cholerae belonging to the recently described serogroup 0139, which are responsible for the current cholera epidemics in India and Bangladesh, were shown to express pilus-like structures partially cross-reacting with the toxin-coregulated pilus of V. cholerae strain (0395) belonging to the 01 serogroup and classical biotype. The 0139 pili were composed of 20 kDa subunit proteins which were antigenically related to the 20 kDa pilus protein of another diarrhoeagenic non-01 V. cholerae strain (serogroup 034) isolated earlier. The pili described in this study were found to be involved in the intestinal colonization process and, therefore, may contribute towards the virulence of the 0139 epidemic isolates.     

Poliovirus-induced intracellular alkalinization involves a proton ATPase and protein phosphorylation

We reported previously that poliovirus infection induces alkalinization in HeLa cells and that an alkaline intracellular pH (pHi) promoted viral replication. Additional experiments were carried out to understand the underlying mechanism. Virus-infected or control monolayer cultures were incubated with nominally bicarbonate-free Eagle's minimal essential medium (MEM) buffered with N-2-hydroxyethylpiperazine-N-3-ethanesulfonic acid (HEPES), and immediately following preincubations, changes in pHi were monitored via benzoic acid uptake around 2 h postinfection. The absence of pH increase in cells infected with ultraviolet light-inactivated virus (UV-virus) indicated that viral gene expression was required for this effect. On the other hand, lack of effect of 3 mM guanidine, an inhibitor of poliovirus-specific RNA but not protein synthesis, suggested that translation of input viral genome RNA is sufficient for the pH increase. Activation of Na⁺/H⁺ exchange, Cl^?HCO^?₃ exchange, or H⁺-ATPase was considered as possible mechanisms by which alkalinization occurs in virus-infected cells. Na⁺/H⁺ exchange was excluded because the pH effect occurred in a Na⁺/H⁺ exchange deficient HeLa cell mutant. Similarly, Cl^?/HCO^?₃ exchange was excluded because virus-specific alkalinization was evident in the presence of Cl^? or bicarbonate deficient medium and was not associated with an increase in HCO^?₃ uptake or a decrease in Cl^? uptake. Lack of dependence on Na⁺, abrogation by 10 μM 7-chloro-4-nitrobenz-2-oxa-1,3-diazole (NBD-Cl), and resistance to 1 mM vandate suggested that this effect was due to the activation of a vacuolar-type (V) proton ATPase. Studies using protein kinase inhibitors indicated that activation of the ATPase in virus-infected cells probably involved protein kinase C-mediated phosphorylation. © 1993 Wiley-Liss, Inc.     

Recognition by and in vitro induction of cytotoxic T lymphocytes against predicted epitopes of the immediate-early protein ICP27 of herpes simplex virus.

The identification of herpes simplex virus type 1 (HSV-1) proteins and the minimal epitopes within these proteins which serve as targets for cytotoxic T lymphocytes (CTL) remains an important goal for the development of effective vaccine strategies. In this report, an H-2Kd allele-specific peptide-binding motif was used to locate putative CTL epitopes in the HSV-1 immediate-early protein ICP27, a protein previously identified as a major CTL target in the BALB/c mouse. Peptides 1 (amino acids 322 to 332) and 2 (amino acids 448 to 456) synthesized to represent two separate predicted CTL epitopes in ICP27 were able to sensitize target cells in vitro for recognition by HSV-1-specific CTL. Moreover, using a recently developed system to generate primary CTL responses in vitro, both peptides induced primary CTL which reacted with target cells expressing HSV-1. This system allowed us to verify the activity of two CTL epitopes in the ICP27 protein and holds promise as a rapid way of identifying immunogenic peptides from any protein molecule.     

A medium for enumeration of bacteria in foods containing swarming Bacillus spp.

A new medium containing sodium tripolyphosphate, ethylenediaminetetraacetic acid disodium salt, dihydrate, sodium desoxycholate, supplemented with minerals and nutrient sources was developed, which was very effective in inhibiting swarming of Bacillus spp. This medium polyphosphate-EDTA-desoxycholate agar (PEDA) was compared with tryptone glucose extract agar (Oxoid) for determining the total plate count of various semi-conserved fishery products. While promoting discrete colony formation of Bacillus spp., PEDA provided good bacterial recovery. PEDA is recommended for isolation or enumeration of bacteria in food dominated by swarming Bacillus spp.