Leishmania donovani adenylate kinase 2a prevents ATP-mediated cell cytolysis in macrophages.

In Leishmania spp. ATP utilizing enzymes serves as a key role in preserving integrity of host cells for survival of parasite. Earlier reports suggested that Adenylate kinase (AK) a phosphotransferase enzyme released by Leishmania donovani secretome, involved in modulating levels of NTPs. In the present study, we cloned, expressed and characterized recombinant putative AK. Based on a sequence and phylogeny analysis, we identified the prominent features of the seven AK isoforms of Leishmania donovani and assigned our putative AK as LdAK2a. The Km value of LdAK2a for ATP and AMP substrate were 204 µM and 184 µM, respectively and Vmax was calculated as 1.6 µmol min-1 mg-1 protein. Ap5A, a known inhibitor of AK inhibited LdAK2a with estimated Ki values of 280 nM and 230 nM for ATP and AMP respectively. CD spectral studies were carried out to estimate its structural stability. Recombinant LdAK2a was found to prevent ATP mediated cell cytolysis of Raw 264.7 macrophages in vitro, which was determined by LDH assay and MMP assay. This is the first report which validates that Leishmanial AK2a can prevent ATP mediated cytolysis of macrophage cells and thereby probably play a role in preserving integrity of host cells for survival of parasite.

Spectrophotometric determination of uranium using ascorbic acid as a chromogenic reagent.

A spectrophotometric method has been developed for the determination of uranium(VI) using ascorbic acid. Uranium in the hexavalent state forms a reddish-brown coloured complex with ascorbic acid. The colour intensity of the complex is maximum at pH 4.2-4.5 and is stable for 24 hr. The absorbances of uranium(VI)-ascorbic acid complex at 360 and 450 nm are used for its quantification. Uranium in the range 8-200 microg/ml has been determined with good precision. The method allows the determination of uranium in the presence of many metal ions present as impurities. The described method is simple, accurate and applicable to uranium concentration relevant to the PUREX process and thus can be used for analytical control purposes.

Inhibition of H+ secretion in frog gastric mucosa by somatostatin.

Somatostatin added to the serosal bathing solution of the isolated gastric mucosa of Rana pipiens significantly inhibited pentagastrin- and histamine-stimulated H+ secretion. The decrease in H+ secretion rate was accompanied by an increase in the transmucosal potential difference and resistance. Somatostatin (10(-5) M) had no effect on the N6,O2-dibutyryl adenosine 3'-5'-cyclic monophosphate (DBcAMP)-stimulated H+ secretion rate. The mucosa exposed to somatostatin secreted H+ on stimulation by DBcAMP or histamine, but did not respond to 2.8 X 10(-7) M pentagastrin. However, pentagastrin added to the serosal solution stimulated H+ secretion after the somatostatin was washed away. Calcium inophore (3 X 10(-5) M) alone or 10(-2) M Ca2+ plus calcium ionophore temporarily increased the H+ secretion rate inhibited by somatostatin. The data suggest that somatostatin has a direct effect on the oxyntic cells in the gastric mucosa.