The use of Fab-masking antigens to enhance the activity of immobilized antibodies

This study demonstrates that masking the Feb regions of a monoclonal antibody (Mab) with synthetic antigens prior to covalent immobilization efficiency. Water-soluble adducts of poly(2-methyloxazoline) polymers and a syntheticpeptide epitope for the Mab were constructed. These synthetic antigens are referred to as Fab-masking antigents (FMAs). The antibody used in this study is a Ca(2+)-dependent murine monoclonal lgG directed against the plasma protein, human protein C (hPC). The FMAs were pre-equilibrated with Mab in the presence of calcium prior to immobilization and were then removed by EDTA, which destabilized the FMA-Mab complexes. The antigen binding efficiency and accessibility of the Fab domain of the immobilized antibody was significantly increased for Mab immobilized in the presence of FMA relative to those Mab immobilized without FMA. The increase in binding efficiency was most pronounced for the largest FMA employed. No appreciable differences were detected in the avidity of hPC-Mab complexes formed by immunosorbents produced by either masked or unmaked antibody. These results provide evidence that orientgation may play an important role in the binding activity of immobilized antibodies.     

Mefenamic Acid Attenuates Chronic Alcohol Induced Cognitive Impairment in Zebrafish: Possible Role of Cholinergic Pathway

Based on the scientific evidence supporting the neuroinflammatory response contributes the cognitive impairment associated with chronic alcoholism and the neuroprotective actions of mefenamic acid with reversal of memory loss and brain inflammation in mice, this study was designed to evaluate the effect of mefenamic acid against chronic alcohol induced cognitive impairment in zebrafish model. Zebrafish were grouped and subjected to normal behavioral analysis in light–dark chamber for 10 days. The preference to dark compartment was noted in zebrafish. Zebrafish were grouped and exposed to escalating doses of alcohol for 28 days with and without mefenamic acid exposure (100 and 200 µg/L) and subjected to a fear conditioning passive avoidance task from day 13 of 28. The cognitive evaluation was performed for 10 days and the brain tissue was isolated to estimate acetylcholinesterase activity. In cognitive evaluation study, the normal zebrafish retained the memory of the learned task and avoided the dark. The alcohol exposed zebrafish showed impairment in retaining the memory of learned task. Mefenamic acid exposed zebrafish showed a significant protection against cognitive impairment caused by alcohol and retained the memory of learned task with a significant decrease in AChE activity in brain homogenate compared to alcohol exposed zebrafish. The results of this study suggest that the memory enhancing activity of mefenamic acid might be due to activation of cholinergic transmission that has protected neuroinflammatory and neurodegenerative conditions caused by alcohol.     

The presence of essential arginine residues at the active sites of citrate lyase complex from Klebsiella aerogenes

The acyl-transferase and acyl-lyase activities of $\text{Klebsiella aerogenes}$ citrate lyase complex are inactivated by the arginine specific reagents phenylglyoxal and 2,3-butanedione, the former reagent being the more potent inhibitor. Citrate and (3$\text{S}$)-citryl-CoA protect the transferase activity, while acetyl-CoA markedly enhances the rate of the inactivation. (3$\text{S}$)-Citryl-CoA protects the lyase subunit in the complex from inactivation. The kinetics of inactivation suggest the involvement of a single arginine residue at each of the active sites of the transferase and of the lyase subunits.     

Toxic and antigrowth effects of raw and processed field bean (Dolichos lablab) on albino rats

Samples of freeze dried green field bean (Dolichos lablab) and dry mature bean, were subjected to the following processing methods—heat processing, extraction with 80% ethanol, hexane or dilute acid, protein isolation; and these samples were evaluated for growth promoting value and toxicity. Extraction with 80% ethanol or with dilute acid increased survival period of the animals; but these did not promote growth. Heat processing was essential to destroy antinutritional factors and promote growth. Extraction of the beans with 80% ethanol did not however alter the trypsin inhibitor or haemagglutinin activities. The protein isolate and acid-extracted residue which had low trypsin inhibitor and haemagglutinin activities, did not also promote growth. Thus the trypsin inhibtor and haemagglutinin activities did not completely account for the toxicity to albino rats. However, heat processing of ethanol extracted bean flour indicated that the beneficial effect of ethanol extraction was not apparent, once the samples were heat processed. Dry mature bean dhal was more toxic than the whole bean either dry or green. Supplementation of heat processed field bean with methionine and tryptophan promoted good growth of albino rats and significantly increased the protein efficiency ratio. Part of the Ph.D. thesis entitled “Studies on the factors affecting the nutritive value of field beanDolichos lablab”, University of Mysore, 1977.     

Three-dimensional structure of the complex of the Rhizopus chinensis carboxyl proteinase and pepstatin at 2.5-A resolution

An X-ray diffraction analysis has been carried out at 2.5-A resolution of the three-dimensional structure of the Rhizopus chinensis carboxyl proteinase complexed with pepstatin. The resulting model of the complex supports the hypothesis [Marciniszyn, J., Hartsuck, J.A., & Tang, J. (1976) J. Biol. Chem. 251, 7088-7094] that statine (3-hydroxy-4-amino-6-methylheptanoic acid) approaches an analogue of the transition state for catalysis. The way in which pepstatin binds to the enzyme can be extended to provide a model of substrate binding and a model of the transition-state complex. This in turn has led to a proposed mechanism of action based on general acid-base catalysis with no covalent intermediates. These predictions are in general agreement with kinetic studies using several carboxyl proteinases, which together with their sequence homology and their common three-dimensional structures suggest that this mechanism can be extrapolated to all carboxyl proteinases.     

A NEW APPROACH TO CORRECTION OF D-LOOP TRANSPOSITION OF THE GREAT ARTERIES

An operative procedure is described for repair of transposition of the great arteries in which the left ventricle is the "aortic" ventricle and the right ventricle is the "pulmonic" ventricle. The technical feasibility of this procedure is supported by its successful application in experimental animals. A modified procedure, which may be applicable in certain complex forms of transposition of the great arteries and other perplexing anomalies for which there are no known repairs, is discussed.     

Association between the p170 form of human topoisomerase II and progeny viral DNA in cells infected with herpes simplex virus type 1.

Endogenous host topoisomerase II acts upon herpes simplex virus type 1 (HSV-1) DNA in infected cells (S.N. Ebert, S.S. Shtrom, and M.T. Muller, J. Virol. 56:4059-4066, 1990), and cleavage is directed exclusively at progeny viral DNA while parental DNA is resistant. To evaluate the possibility that HSV-1 induces topoisomerase II activity which could account for the preferential cleavage of progeny viral DNA, we assessed topoisomerase II cleavage activity on cellular and viral DNA substrates before and after the initiation of viral DNA replication. We show that cleavage of a host gene in mock-infected cells was similar to that observed in HSV-1-infected cells, regardless of whether viral DNA replication had occurred. In addition, quantitative measurements revealed comparable amounts of topoisomerase II activity in infected and mock-infected cells; thus, HSV-1 neither induces nor encodes its own type II topoisomerase and cleavages in vivo are due to a preexisting host topoisomerase. Human cells contain two isozymes of topoisomerase II (p170 and p180), encoded by separate genes. Through the use of isozyme-specific antibodies, we demonstrate that only p170 was found to be cross-linked to HSV-1 DNA even though both forms were present at nearly constant levels in HSV-1-infected cells. Immunofluorescence revealed that by 6 h postinfection, p170 becomes redistributed and localized to sites of active viral DNA synthesis. The data suggest that p170 gains preferential access to replicated viral DNA molecules, which explains why topoisomerase II activity is concentrated on progeny DNA.     

Panchanania andPhragmospathula,two new genera of the Hyphomycetes

Two new Hyphomycetes collected from Jaipur, Rajasthan, India are described.Panchanania jaipurensis gen. et sp.n., collected on dead and dying twigs ofGrewia salvifolia has characteristic solitary stalked blastospores, each with a dark-coloured head and paler stalk, on sympodulae borne on synnemata.Phragmospathula phoenicis gen. et sp.n., collected on dead leaf rachis ofPhoenix sp. has characteristic solitary spathulate phragmospores (blastospores) which are dark-coloured in the middle and paler at the ends and are produced on short conidiophores capable of proliferation from within. On the basis of the mode of development of the spores, both fungi are placed in the Torulaceae Corda emend. Subram.