Cloning, expression, characterization, and inhibition studies on Trypanothione Synthetase, a drug target enzyme, fromLeishmania donovani

2011 ◽  
pp. ---
Author(s):  
Prakash Saudagar, ◽  
Vikash Kumar Dubey,
Keyword(s):  
Drug Target ◽  
Expression Characterization ◽  
Inhibition Studies ◽  
Target Enzyme

Cloning, expression, characterization and inhibition studies on trypanothione synthetase, a drug target enzyme, from Leishmania donovani

2011 ◽  
Vol 392 (12) ◽  
pp. 1113-1122 ◽  
Author(s):  
Prakash Saudagar ◽  
Vikash Kumar Dubey
Keyword(s):  
Gene Cloning ◽  
Drug Target ◽  
Leishmania Donovani ◽  
Redox Homeostasis ◽  
Substrate Inhibition ◽  
Leishmanicidal Activity ◽  
High Substrate ◽  
Glutathione Concentration ◽  
Expression Characterization ◽  
Inhibition Studies

Abstract Trypanothione synthetase, a validated drug target, synthesizes trypanothione from glutathione and spermidine. Here we report the gene cloning, expression, characterization and inhibition studies of trypanothione synthetase from Leishmania donovani (LdTryS). The purified recombinant LdTryS enzyme obeyed Michaelis-Menten kinetics. High substrate inhibition was observed with glutathione (Km=33.24 μm, kcat=1.3 s-1, Ki=866 μm). The enzyme shows simple hyperbolic kinetics with fixed glutathione concentration and with other substrates limiting Km values for Mg. ATP and spermidine of 14.2 μm and 139.6 μm, respectively. LdTryS was also screened for inhibitors. Tomatine, conessine, uvaol and betulin were identified as inhibitors of the enzyme and were tested for leishmanicidal activity. Finally, the effect of LdTryS inhibitors on redox homeostasis of the parasite gives a broader picture of their action against leishmaniasis.

Haplotype structures and functional polymorphic variants of the drug target enzyme aromatase (CYP19A1) in South Indian population

2013 ◽  
Vol 30 (3) ◽  
Author(s):  
Gurusamy Umamaheswaran ◽  
Steven Aibor Dkhar ◽  
Sekar Kalaivani ◽  
Raj Anjana ◽  
Mohan Revathy ◽  
...  
Keyword(s):  
Drug Target ◽  
Indian Population ◽  
South Indian Population ◽  
South Indian ◽  
Polymorphic Variants ◽  
Target Enzyme

scholarly journals Expression of a recombinant, 4’-Phosphopantetheinylated, active M. tuberculosis Fatty acid Synthase I in E. coli

2018 ◽  
Author(s):  
Szilvia Baron ◽  
Yoav Peleg ◽  
Jacob Grunwald ◽  
David Morgenstern ◽  
Nadav Elad ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Drug Target ◽  
Fatty Acid Synthase ◽  
Acyl Carrier Protein ◽  
Spectrophotometric Analysis ◽  
Carrier Protein ◽  
E Coli ◽  
Drug Inhibition ◽  
Nadph Oxidation ◽  
Inhibition Studies

AbstractFatty acid synthase 1 (FAS I) from Mycobacterium. tuberculosis (Mtb) is an essential protein and a promising drug target. FAS I is a multi-functional, multi-domain protein that is organized as a large (1.9 MDa) homohexameric complex. Acyl intermediates produced during fatty acid elongation are attached covalently to an acyl carrier protein (ACP) domain. This domain is activated by the transfer of a 4’-Phosphopantetheine (4’-PP, also termed P-pant) group from CoA to ACP catalyzed by a 4’-PP transferase, termed acyl carrier protein synthase (AcpS). In order to obtain an activated FAS I in E. coli, we transformed E. coli with tagged Mtb fas1 and acpS genes encoded by a separate plasmid.We induced the expression of Mtb FAS I following induction of AcpS expression. FAS I was purified by Strep-Tactin affinity chromatography. Activation of Mtb FAS I was confirmed by the identification of a bound P-pant group on serine at position 1808 by mass spectrometry. The purified FAS I displayed biochemical activity shown by spectrophotometric analysis of NADPH oxidation and by CoA production, using the Ellman reaction. The purified Mtb FAS I forms a hexameric complex shown by negative staining and cryo-EM. Purified hexameric and active Mtb FAS I is required for binding and drug inhibition studies and for structurefunction analysis of this enzyme. This relatively simple and short procedure for Mtb FAS I production should facilitate studies of this enzyme.

scholarly journals Curcumin binds in silico to anti-cancer drug target enzyme MMP-3 (human stromelysin-1) with affinity comparable to two known inhibitors of the enzyme

2015 ◽  
Vol 11 (8) ◽  
pp. 387-392 ◽  
Author(s):  
Ahmed Jerah ◽  
◽  
Yahya Hobani ◽  
Vinod Kumar ◽  
Anil Bidwai ◽  
...  
Keyword(s):  
In Silico ◽  
Drug Target ◽  
Cancer Drug ◽  
Anti Cancer ◽  
Target Enzyme

scholarly journals Sulfonamide Inhibition Studies of an α-Carbonic Anhydrase from Schistosoma mansoni, a Platyhelminth Parasite Responsible for Schistosomiasis

2020 ◽  
Vol 21 (5) ◽  
pp. 1842 ◽  
Author(s):  
Andrea Angeli ◽  
Mariana Pinteala ◽  
Stelian S. Maier ◽  
Bogdan C. Simionescu ◽  
Akram A. Da’dara ◽  
...  
Keyword(s):  
Carbonic Anhydrase ◽  
Schistosoma Mansoni ◽  
Drug Target ◽  
Hydration Reaction ◽  
High Catalytic Activity ◽  
Intestinal Schistosomiasis ◽  
Host Parasite ◽  
Inhibition Studies ◽  
Platyhelminth Parasite ◽  
Aromatic Heterocyclic

Schistosomiasis is a debilitating infection provoked by parasitic flatworms called schistosomes. The species Schistosoma mansoni is endemic in Africa, where it causes intestinal schistosomiasis. Recently, an α-carbonic anhydrase (CA, EC 4.2.1.1) was cloned and characterized from this organism and designated as SmCA. The protein is expressed in the tegument (skin) of S. mansoni at the host–parasite interface. Recombinant SmCA possesses high catalytic activity in the CO2 hydration reaction, similar to that of human CA isoform II with a kcat of 1.2 × 106 s−1 and a kcat/KM of 1.3 × 108 M−1·s−1. It has been found that schistosomes whose SmCA gene is suppressed using RNA interference are unable to establish a robust infection in mice, suggesting that the chemicals that inhibit SmCA function should have the same debilitating effect on the parasites. In this study, a collection of aromatic/heterocyclic sulfonamides were investigated as possible SmCA inhibitors. Several sulfonamides inhibited SmCA with medium to weak potency (KI values of 737.2 nM−9.25 μM), whereas some heterocyclic compounds inhibited the enzyme with KI values in the range of 124−325 nM. The α-CA from S. mansoni, SmCA, is proposed as a new anti-schistosomiasis drug target.

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