Interaction of a fluorescent analog of N -deacetyl-N -methyl-colchicine (colcemid) with liver alcohol dehydrogenase

The evidence for specific binding of N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-colcemid (NBD-colcemid), a fluorescent analog of colcemid (N-deacetyl-N-methyl-colchicine), to liver alcohol dehydrogenase is presented. Alcohol dehydrogenase bound NBD-colcemid in a time-dependent manner, enhanced the fluorescence intensity, and caused a large blue shift of the emission maximum of the free drug. The specificity of binding was determined for both the colchicine nucleus and the NBD moiety. The binding was not affected by the presence of alcohol or NAD in the reaction mixture. Preincubation of horse liver alcohol dehydrogenase with colcemid inhibited the binding to a considerable extent. NBD-colcemid inhibited the enzymic activity of alcohol dehydrogenase in a mixed-type noncompetitive mode with a Ki value of 32 μM, whereas colcemid showed noncompetitive inhibition with a Ki of 100 μM. The association rate constant of NBD-colcemid binding with liver alcohol dehydrogenase was 587 M-1 s-1 at 25 °C. The stoichiometry and dissociation constant of the binding reaction were 0.62/dimer and 12 μM, respectively. Donor quenching experiments showed that both tryptophans of alcohol dehydrogenase transferred energy to the bound NBD-colcemid. Thus, this study reports the binding of a colchicine analog to a protein other than tubulin with high affinity. It is concluded that NBD-colcemid binding to dehydrogenases is a general phenomenon, but the common structural element(s) that is responsible for the binding activity, and which exists among tubulin and dehydrogenases, has yet to be determined

Drug Trafficking Routes and Hepatitis B in Injection Drug Users, Manipur, India

Prevalence of hepatitis B genotype C in injection drug users in the northeastern Indian state of Manipur, neighboring the "Golden Triangle," correlates well with overland drug-trafficking routes, the injection drug use epidemic, and the spread of HIV. Further spread to other regions of India through mobile populations is possible

Unveiling the nongenomic actions of thyroid hormones in adult mammalian brain: The legacy of Mary B. Dratman

A comprehensive review was conducted to compile the contributions of Mary B. Dratman and studies by other researchers in the field of nongenomic actions of thyroid hormones in adult mammalian brain. Dratman and her collaborators authored roughly half of the papers in this area. It has been almost fifty years since Dratman introduced the novel concept of thyroid hormones as neurotransmitters for the first time. The characterization of unique brain-region specific accumulation of thyroid hormones within the nerve terminals in adult mammals was a remarkable contribution by Dratman. It suggested a neurotransmitter- or neuromodulator-like role of thyroid hormone and/or its derivative, 3-iodothyronamine within adrenergic systems in adult mammalian brain. Several studies by other researchers using synaptosomes as a model system, have contributed to the concept of direct nongenomic actions of thyroid hormones at synaptic regions by establishing that thyroid hormones or their derivatives can bind to synaptosomal membranes, alter membrane functions including enzymatic activities and ion transport, elicit Ca2+/NO-dependent signaling pathways and induce substrate-protein phosphorylation. Such findings can help to explain the physiological and pathophysiological roles of thyroid hormone in psychobehavioral control in adult mammalian brain. However, the exact mode of nongenomic actions of thyroid hormones at nerve terminals in adult mammalian brain awaits further study

Anti-hepatitis B core antigen testing with detection and characterization of occult hepatitis B virus by an in-house nucleic acid testing among blood donors in Behrampur, Ganjam, Orissa in southeastern India: implications for transfusion

<p>Abstract</p> <p>Background</p> <p>Occult hepatitis B virus (HBV) infection might transmit viremic units into the public blood supply if only hepatitis B surface antigen (HBsAg) testing is used for donor screening. Our aim was to evaluate the prevalence of occult HBV infection among the HBsAg negative/antiHBc positive donations from a highly HIV prevalent region of India.</p> <p>Methods</p> <p>A total of 729 HBsAg negative donor units were included in this study. Surface gene and precore region were amplified by in house nucleic acid test (NAT) for detection of occult HBV infection and surface gene was analyzed after direct sequencing.</p> <p>Results</p> <p>A total of 220 (30.1%) HBsAg negative donors were antiHBc positive, of them 66 (30%) were HBV DNA positive by NAT. HBV DNA positivity among 164 antiHBc only group, was 27.1% and among 40 antiHBs positive group was 30.0%. HBV/D (93.3%) was predominant and prevalence of both HBV/C and HBV/A was 3.3%. Single or multiple amino acids substitutions were found in 95% samples.</p> <p>Conclusion</p> <p>Thus, a considerable number of HBV infected donors remain undiagnosed, if only HBsAg is used for screening. Addition of antiHBc testing for donor screening, although will lead to rejection of a large number of donor units, will definitely eliminate HBV infected donations and help in reducing HBV transmission with its potential consequences, especially among the immunocompromised population. The HBV genetic diversity found in this donor population are in accordance with other parts of India.</p

Simultaneous administration of adjuvant donor bone marrow in pancreas transplant recipients

Objective: The effect of donor bone marrow was evaluated for its potentially favorable effect in the authors' simultaneous pancreas/kidney transplant program. Methods: From July 1994 to January 1999, 177 pancreas transplants were performed, 151 of which were simultaneous pancreas/kidney transplants. All patients received tacrolimus, mycophenolate mofetil, and steroids for immunosuppression (azathioprine was used in the first year of the program). Fifty-three simultaneous pancreas/kidney transplant recipients received perioperative unmodified donor bone marrow, 3 to 6 x 108 cells/kg. Results: Overall actuarial survival rates at 1 and 3 years were 98% and 95% (patient), 95% and 87% (kidney), and 86% and 80% (pancreas), respectively. In the adjuvant bone marrow group, 1- and 3-year survival rates were 96% and 91% (patient), 95% and 87% (kidney), and 83% and 83% (pancreas), respectively. For 98 recipients who did not receive bone marrow, survival rates at 1 and 3 years were 100% and 98% (patient), 96% and 86% (kidney), and 87% and 79% (pancreas), respectively. No pancreas allografts were lost after 3 months in bone marrow recipients, and seven in the non-bone marrow recipients were lost to rejection at 0.7, 6.7, 8.8, 14.6, 24.1, 24.3, and 25.5 months. Twenty-two percent of bone marrow patients were steroid-free at 1 year, 45% at 2 years, and 67% at 3 years. Nineteen percent of the non-bone marrow recipients were steroid-free at 1 year, 38% at 2 years, and 45% (p = 0.02) at 3 years. The mean acute cellular rejection rate was 0.94 ± 1.1 in the bone marrow group and 1.57 ± 1.3 (p = 0.003) in the non-bone marrow group (includes borderline rejection and multiple rejections). The level of donor cell chimerism in the peripheral blood of bone marrow patients was at least two logs higher than in controls. Conclusion: In this series, which represents the largest experience with adjuvant bone marrow infusion in pancreas recipients, there was a higher steroid withdrawal rate (p = 0.02), fewer rejection episodes, and no pancreas graft loss after 3 months in bone marrow recipients compared with contemporaneous controls. All pancreas allografts lost to chronic rejection (n = 6) were in the non-bone marrow group. Donor bone marrow administered around the time of surgery may have a protective effect in pancreas transplantation

Globalisation of technology development: offshoring of R&D to China, India and Brazil

Fundamental shifts in the mode, the geography and the personnel of global technology development are taking place in the wake of increasing globalisation of economies and rapid technological changes. Many large multinational corporations are focusing on building global innovation networks, comprising of R&D units located in clusters of excellence around the world, including many developing economies. In this paper we take a close look at this emerging phenomenon – particularly the developments taking place in China, India and Brazil. By using US patents as indicators, this study seeks to detect the trends in the technical activities, the innovative capability, the ownership of the intellectual property and the quality of technology developed in these countries.globalisation; offshoring; R&D; research and development; China; India; Brazil; patents; science and technology; technology development; multinational corporations; MNCs; global innovation networks; industry clusters; developing countries; innovative capability; intellectual property ownership; technology quality; innovation; technological change.

Amino acid residues involved in autophosphorylation and phosphotransfer activities are distinct in nucleoside diphosphate kinase from mycobacterium tuberculosis

Nucleoside diphosphate kinase (NdK) is a ubiquitous enzyme in both prokaryotes and eukaryotes and is primarily involved in the maintenance of cellular nucleotide pools. We have cloned ndk from Mycobacterium tuberculosis strain H37Ra and expressed it in Escherichia coli as a fusion protein with glutathione S-transferase. The purified protein, following thrombin cleavage and gel permeation chromatography, was found to be hexameric with a monomeric unit molecular mass of ~16.5 kDa. The protein exhibited nucleotide binding, divalent cation-dependent autophosphorylation, and phosphate transfer ability from nucleoside triphosphate to nucleoside diphosphate. Although UDP inhibited the catalytic activity of the recombinant protein, the classic inhibitors, like cromoglycate, 5'-adenosine 3'-phosphate, and adenosine 3'-phosphate 5'-phosphosulfate, had no effect on the activity. Among three histidine residues in the protein, His-117 was found to be essential for autophosphorylation. However, in subsequent phosphate transfer, we observed that His-53 had a significant contribution. Consistent with this observation, substitution of His-53 with either Ala or Gln affected the ability of the recombinant protein to complement NdK function in Pseudomonas aeruginosa. Furthermore, mutational analysis established critical roles for Tyr-50 and Arg-86 of the M. tuberculosis protein in maintaining phosphotransfer ability