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Biomedical subjects

B Bhattacharyya

Publications and source records attributed to B Bhattacharyya.

At least 55 records · Page 3Linked to original sources
PubMed1992

Multiphasic denaturation of the lambda repressor by urea and its implications for the repressor structure.

Urea denaturation of the lambda repressor has been studied by fluorescence and circular dichroic spectroscopies. Three phases of denaturation could be detected which we have assigned to part of the C-terminal domain, N-terminal domain and subunit dissociation coupled with further denaturation of the rest of the C-terminal domain at increasing urea concentrations. Acrylamide quenching suggests that at least one of the three tryptophan residues of the lambda repressor is in a different environment and its emission maximum is considerably blue-shifted. The transition in low urea concentration (midpoint approximately 2 M) affects the environment of this tryptophan residue, which is located in the C-terminal domain. Removal of the hinge and the N-terminal domain shifts this transition towards even lower urea concentrations, indicating the presence of interaction between hinge on N-terminal and C-terminal domains in the intact repressor.

DNA-Binding Proteins↗
PubMed1992

An operator-induced conformational change in the C-terminal domain of the lambda repressor.

4,4'-bis(1-anilino-8-naphthalenesulfonic acid (Bis-ANS), an environment-sensitive fluorescent probe for hydrophobic region of proteins, binds specifically to the C-terminal domain of lambda repressor. The binding is characterized by positive cooperativity, the magnitude of which is dependent on protein concentration in the concentration range where dimeric repressor aggregates to a tetramer. In this range, positive cooperativity becomes more pronounced at higher protein concentrations. This suggests a preferential binding of Bis-ANS to the dimeric form of the repressor. Binding of single operator OR1 to the N-terminal domain of the repressor causes enhancement of fluorescence of the C-terminal domain bound Bis-ANS. The binding of single operator OR1 also leads to quenching of fluorescence of tryptophan residues, all of which are located in the hinge or the C-terminal domain. Thus two different fluorescent probes indicate an operator-induced conformational change which affects the C-terminal domain. The significance of this conformational change with respect to the function of lambda repressor has been discussed.

Anilino Naphthalenesulfonates↗
PubMed1992

Excimer fluorescence of pyrene-maleimide-labeled tubulin.

Excimer-forming cysteines in tubulin are detected by the presence of excimer fluorescence in N-(1-pyrenyl)maleimide-labeled tubulin. The ratio of excimer/monomer fluorescence of labeled protein remained unchanged upon its dilution. These results indicating that both partner of each pair(s) of cysteine are located in the same subunit. The excimer fluorescence is insensitive to prior treatment of tubulin with either colchicine or GTP, indicating that pairs of cysteines protected by those drugs are not involved in excimer formation. This excimer fluorescence of N-(1-pyrenyl)maleimide-labeled tubulin disappeared upon treatment with SDS, guanidinium chloride (GdmCl) and urea. Studies with GdmCl induced unfolding of N-(1-pyrenyl)maleimide-labeled tubulin showed that the loss of excimer fluorescence precedes subunit dissociation. The loss of both colchicine-binding activity and the excimer fluorescence with increasing temperature indicates a major conformational change of the tubulin molecule at elevated temperatures.

Guanidine↗
PubMed1992

Interaction of Prodan with tubulin. A fluorescence spectroscopic study.

The compound 6-propionyl-2-(N,N-dimethyl)-aminonaphthalene (Prodan), an efficient fluorescent probe for proteins, is shown to bind to tubulin. Detailed experiments on fluorescence enhancement, anisotropy and energy transfer were carried out to unravel the nature of Prodan-tubulin interaction and the Prodan-binding site on tubulin. It was found that Prodan binds to tubulin at a rigid site, with a stoichiometry of 1:1 and a dissociation constant of 20 microM. Competition experiments using 1,8-[3H]anilinonaphthalenesulfonate ([3H]ANS) indicated that the three aminonaphthalenes, ANS, bis(1,8-anilinonaphthalenesulfonate) and Prodan probably bind to a common region on the tubulin molecule.

2-Naphthylamine↗
PubMed1992

A retrospective study on scorpion sting in a pediatric age group in a hospital in Calcutta.

During 1985-1989, in Calcutta Medical College Hospitals, of 152 children of 1-6 year age group admitted with the history of scorpion sting 18 (11.8%) died. Maximum numbers of stings were inflicted in the fingers. Important clinical features recorded were circulatory failure, breathlessness, profuse sweating, vomiting, local oedema and convulsion. Incidences of scorpion stings were much more frequent in the summer and rainy seasons than in the winter season.

Animals↗
PubMed1990

The carboxy terminus of the alpha subunit of tubulin regulates its interaction with colchicine.

Controlled proteolysis of goat brain tubulin by subtilisin was carried out to investigate regulatory aspects of the binding of colchicine to tubulin. Tubulin S, obtained by the cleavage of the carboxyl termini of both the alpha- and beta-subunits of tubulin by subtilisin, exhibited the following differences compared to native tubulin: (a) Reaction with colchicine, which has an optimum pH of 6.8, becomes independent of pH (in the range 5.7-8.0). (b) The colchicine-binding site, which is labile at 37 degrees C (t1/2 = 4-5 h), becomes highly stable (t1/2 greater than 12 h). (c) The affinity for colchicine is lowered. (d) This lowering of affinity arises from a faster dissociation (higher off rate) of the complex. The above characteristics of tubulin S were not shown by a partially digested hybrid in which the C-terminus of the beta-subunit alone was cleaved. The hybrid behaved very much like the undigested native protein. These results strongly suggest that the regulatory switch for colchicine-tubulin interaction is located in a small region (about 15 residues) of the C-terminus of the alpha-subunit of tubulin. Possibilities of the C-termini being involved in nonbonded contacts with the main body of tubulin are also noticed from the change in conformation between tubulin and tubulin S.

Animals↗
PubMed1989

Halothane and isoflurane alter acetylcholine activated ion channel kinetics.

The effects of halothane and isoflurane on the acetylcholine activated ion channel were studied in the frog sartorius muscle using the two electrode voltage clamp technique. The miniature end-plate currents (MEPCs) were recorded and evaluated for amplitude, duration of growth phase and time constant of decay (tau). Both halothane and isoflurane decreased tau in a dose dependent manner. Depression of current amplitude was also dose dependent. The ED50 value indicates tau is more affected than the amplitude of MEPC. Biexponential decay phases were seen in a small fraction of cells exposed to a low concentration of halothane but not isoflurane. Biexponential decay, when seen, was more prominent at less negative clamped membrane potential. The normal linear relationship between membrane potential and tau was not altered by anesthetics.

Acetylcholine↗
PubMed1989

Use of micelles in studying drug-binding sites: simulation of the tubulin-bound fluorescence of colchicine.

Enhancement of the fluorescence intensity of colchicine occurs in media of low polarity and appreciable viscosity; this is suggested to be the basis of the intensification of its fluorescence when it is bound to and immobilized in tubulin. We show here that the tubulin-bound fluorescence features of colchicine are largely reconstructed upon solubilizing it in chosen micellar aggregates that offer optimal polarities and microviscosities. Triton X-100 and bile salt micelles intensify the colchicine emission but the maximal effects are obtained with tetrameric aggregates of the peptide melittin. Estimates of the polarity, microviscosity and binding-site dimensions of colchicine are obtained using this mimetic approach. Our results suggest that well chosen micellar systems act as good models to reconstruct and analyze the spectral properties of molecules immobilized in their binding sites.

Binding Sites↗
PubMed1989

Isolation, characterization and biological activities of a toxin from Bacillus megaterium (B-23).

Fungitoxic substance was isolated from the culture filtrate of B. megaterium (B-23). Age of culture and pH of medium influence the fungitoxicity of its culture filtrate. Partially purified toxin was thermolabile, non-dialysable, ethyl acetate soluble, vanillin-sulphuric acid positive and effective within a range of pH 5-9. It exhibited maximum UV absorption at 224 nm. Its melting point was 242 degrees C. The efficacy of this compound was tested on 4 jute parasites namely, C. corchori, C. gloeosporioides, M. roridum and A. citri, of which M. roridum and C. corchori were least and most sensitive to the toxin respectively.

Bacillus megaterium↗
PubMed1988

Neuromuscular blocking action of two hemicholinium-3 analogs.

Two hemicholinium-3 (HC-3) analogs (NAM-204 and NAM-224) in which the C = O moiety was reduced to -CH2 and 2- or 3-CH3 substitution on the piperidine ring were demonstrated to produce a different type of neuromuscular inhibition than that induced by HC-3. The neuromuscular blocking activity was tested in several different preparations and it was shown that these compounds are potent, short-acting, and was reversed by neostigmine but not by choline. Although these compounds possess weak anticholinesterase activity, significant alterations of blood pressure and heart rate are absent. Miniature end-plate potentials' amplitude were found to be diminished by these compounds with some alteration of frequency. In voltage-clamped frog sartorius muscle NAM-224 and NAM-204 decreased amplitude of miniature end-plate current (mepcs) and accelerated the time constant of mepc decay in a concentration-dependent manner, without altering the single exponential nature of mepc decay. NAM-204 and NAM-224 at high concentrations also produced a voltage- and concentration-dependent non-linearity in current/voltage relationships, especially at more negative membrane potentials. Hence, these pharmacological and electrophysiological studies indicate NAM-224 and NAM-204 have significant activity at both pre- and post-junctional sites and their post-junctional blocking activity results from blocking ACh receptor/ion channel complex.

Action Potentials↗
PubMed1988

Structure-activity relationship studies of hemicholinium (HC-3) congeners.

In a continuing investigation of structural requirements for hemicholinium-like activity (inhibition of neuromuscular transmission due to inhibition of uptake of choline into nerve terminals), some additional molecular modifications of hemicholinium ("HC-3"; structure 1) were made. The target compounds were prepared by standard one- or two-step sequences. Noncyclic acetal moieties in general permitted retention of pharmacological activity, as did concomitant replacement of the central biphenyl "spacer" by other bulky cyclic groupings and replacement of the oxazinium rings by piperidine or 4-methylpiperidine. However, these modifications generally produced compounds of a lower potency. Replacement of the biphenyl moiety of HC-3 with polyakylene chains permitted retention of a considerable degree of activity. In these target compounds, the two quaternary nitrogens can exist the same distance apart (approximately 14 A) as in the hemicholinium molecule. The ditertiary amino congener of a pharmacologically active bis-quaternary oxazinium compound was almost completely inactive. To date, only one tertiary amine has been found which displays a significant degree of hemicholinium-like activity.

Animals↗
PubMed1987

Properties of B-ring analogues of colchicine.

Absorption spectra of colchicine and its analogues are affected by the presence of the B-ring, although it is not part of the chromophore (C-ring). Thus, 2-methoxy-5-(2',3'4'-trimethoxyphenyl)tropone has absorption maxima at 341 nm, whereas that of desacetamidocolchicine is at 353 nm. A similar red shift in the lambda max of colchicine, desacetamidocolchicine and 2-methoxy-(2',3',4'-trimethoxyphenyl)tropone also occurs when they are immobilized in the binding site to tubulin or in pure glycerol. We also observed that the B-ring of colchicine alone or with substituent does not affect the UV-induced rearrangement of colchicine to lumicolchicine. However, in the absence of the B-ring, as in the case of 2-methoxy-5-(2',3'4'-trimethoxyphenyl)tropone, the rearrangement reaction of the C-ring slows down significantly.

Colchicine↗
PubMed1987

Kinetic and thermodynamic analysis of taxol-induced polymerization of purified tubulin.

The kinetic and thermodynamic behavior of in vitro taxol-induced polymerization of purified tubulin has been studied. The assembly of tubulin initiated by taxol has a critical concentration of 0.1 mg/mL at 37 degrees C and consists of two consecutive pseudo first-order processes, a fast phase followed by a slow phase. The rate constants of the fast and slow phase polymerizations increase linearly with increasing tubulin concentration. This implies that the polymerization is a true pseudo first-order process. The ln (l/t0.5) of polymerization for both fast and slow phases follows a linear function with ln [tubulin] fulfilling one of the criteria of condensation polymerization mechanism. From the Arrhenius plot, the temperature dependence of the rate of tubulin polymerization in the presence of taxol is biphasic. The apparent activation enthalpies for the overall polymerization reaction are 13.0 and 50.8 kcal/mol (1 cal = 4.1868 J), respectively, above and below 26 degrees C. The apparent activation enthalpies for the elongation reaction have also been determined. The values are 11.6 and 28.4 kcal/mol above and below 28 degrees C. The temperature dependence of the equilibrium constants as revealed by the van't Hoff plot is also biphasic. The standard enthalpy and entropy values are delta H degrees = 7.4 and 22.5 kcal/mol above and below 30 degrees C, and delta S degrees = 50.3 and 101.0 cal/(deg.mol), at high and low temperatures, respectively. This suggests that the taxol-induced assembly of purified tubulin is a process driven by the effect of entropy.

Alkaloids↗
PubMed1987

Pharmacologic evaluation and structure activity relationships of a series of hemicholinium-3 (HC-3) analogs.

Piperidine derivatives of hemicholinium-3 were synthesized, which included the following spacing groups between cationic heads: trans-trans-bicyclohexyl, phenanthrene, naphthalene, and biphenyl. Relatively minor structural alterations in these series of compounds resulted in several different types of pharmacological actions related to cholinergic transmission. Structural requirements of the compounds are discussed and include internitrogen distance, structural planarity, spacing groups, and positional isomerism of the quaternary cationic heads. Selected quaternary piperidine derivatives with 14 A inter-atomic distance between the cationic heads exhibit potent HC-3 like activity which is enhanced if a molecule has a nonpolar space filling group (4-methyl piperidine) approximately 3.7 A from the corresponding quaternary cationic head. With selected piperidine ring substitutions, active tertiary amines were also identified. Compounds containing C = O in the spacing moiety were active inhibitors of cholinesterase with some derivatives being nearly as active as physostigmine. When the C = O moiety was reduced to -CH2 and a 2 or 3-CH3 piperidine ring was present, potent non-depolarizing, short acting neuromuscular blocking agents were obtained.

Animals↗
PubMed1987

Structure-activity relationship studies in the hemicholinium ('HC-3') series.

A series of congeners of hemicholinium ('HC-3') has been synthesized, in which the central biphenyl portion of the molecule has been modified, in order to gain insight into possible structural requirements of the biphenyl moiety needed for inhibition of choline uptake into nerve terminals. All molecular modifications resulted in compounds with qualitatively and quantitatively similar pharmacological properties to hemicholinium. It is concluded that the biphenyl portion of the hemicholinium molecule serves as a spacer for maintaining the separation of either the quaternary nitrogens, or the oxazonium rings, at an optimal distance for appropriate interaction with receptor site(s).

Animals↗
PubMed1986

Anticholinesterase activity and structure activity relationships of a new series of hemicholinium-3 analogs.

Piperidine derivatives of hemicholinium-3 were synthesized and included the following spacing groups between the bis-cationic heads: trans,trans-bicyclohexyl, phenanthrene, naphthalene and biphenyl. Anticholinesterase activity was determined using rat striatal synaptosomal cholinesterase, bovine erythrocyte acetylcholinesterase and horse serum butyrylcholinesterase. Hemicholinium-3 has little anticholinesterase activity but when the choline moiety of hemicholinium-3 is replaced with selected heterocyclic amine ring systems active inhibitors can be obtained. Results in this communication identify several quaternary amines which are equipotent with physostigmine for inhibition of cholinesterase. Several tertiary amines were also found to be active. Optimal anticholinesterase activity of these piperidine derivatives appear to be related to the necessity of 14 A interatomic distance between the cationic heads as well as C = O substitution in the phenylethyl spacing moiety. Reduction of C = O to secondary alcohol or CH2 results in decreased activity. The anticholinesterase activity is not only related to internitrogen distance and C = O substitution but also structural planarity and positional isomerism of the quaternary cationic head.

Animals↗
PubMed1986

Evaluation of 4-methylpiperidine analogs of hemicholinium-3.

A series of substituted piperidine analogs of hemicholinium-3 was evaluated for their ability to inhibit neuromuscular transmission, to decrease acetylcholine content of caudate slices, to inhibit choline acetyltransferase activity, and to produce toxicity. Quaternary and tertiary amine derivatives of 4-methyl- and 4-hydroxyl-substituted piperidine analogs containing beta-carbonyl or beta-hydroxyl substitutions in the phenylethyl spacing moiety were tested. 4-Methyl piperidine derivatives maintained potent hemicholinium-3 like activity. Reduction of activity was seen with the 4-hydroxyl piperidine analogs. Compounds with beta-hydroxyl substitution were more potent than those with beta-carbonyl substitution. The tertiary amine, 4-methyl piperidine derivative with a hydroxyl group on the beta-carbon of the ethyl side chain also possessed hemicholinium-3 like activity. However, tertiary amine analogs were substantially less potent than hemicholinium-3 or their quaternary amine analogs.

Acetylcholine↗