[show abstract][hide abstract] ABSTRACT: We have determined the crystal structure of porcine quinolinate phosphoribosyltransferase (QAPRTase) in complex with nicotinate mononucleotide (NAMN), which is the first crystal structure of a mammalian QAPRTase with its reaction product. The structure was determined from protein obtained from the porcine kidney. Because the full protein sequence of porcine QAPRTase was not available in either protein or nucleotide databases, cDNA was synthesized using reverse transcriptase-polymerase chain reaction to determine the porcine QAPRTase amino acid sequence. The crystal structure revealed that porcine QAPRTases have a hexameric structure that is similar to other eukaryotic QAPRTases, such as the human and yeast enzymes. However, the interaction between NAMN and porcine QAPRTase was different from the interaction found in prokaryotic enzymes, such as those of Helicobacter pylori and Mycobacterium tuberculosis. The crystal structure of porcine QAPRTase in complex with NAMN provides a structural framework for understanding the unique properties of the mammalian QAPRTase active site and designing new antibiotics that are selective for the QAPRTases of pathogenic bacteria, such as H. pylori and M. tuberculosis.
PLoS ONE 01/2013; 8(4):e62027. · 3.73 Impact Factor
[show abstract][hide abstract] ABSTRACT: As previously reported, the activity of the large-conductance calcium (Ca(2+))-activated potassium (K(+)) (BK(Ca)) channel is strongly potentiated from the extracellular side of the cell membrane by certain benzofuroindole derivatives. Here, the mechanism of action of one of the most potent activators, 4-chloro-7-(trifluoromethyl)-10H-benzofuro[3,2-b]indole-1-carboxylic acid (CTBIC), is characterized. This compound, Compound 22 in the previous report (Chembiochem 6:1745-1748, 2005), potentiated the activity of the channel by shifting its conductance-voltage relationship toward the more negative direction. Cotreatment with CTBIC reduced the affinity of charybdotoxin, a peptide pore-blocker, whereas that of tetraethylammonium, a small pore-blocking quaternary ammonium, was not significantly altered. Guided by these results, scanning mutagenesis of the outer vestibule of the BK(Ca) channel was launched to uncover the molecular determinants that affect CTBIC binding. Alanine substitution of several amino acid residues in the turret region and the S6 helix of the channel decreased potentiation by CTBIC. Homology modeling and molecular dynamics simulation showed that some of these residues formed a CTBIC binding pocket between two adjacent α-subunits in the outer vestibule of the channel. Thus, it can be envisioned that benzofuroindole derivatives stabilize the open conformation of the channel by binding to the residues clustered across the extracellular part of the subunit interface. The present results indicate that the interface between different α-subunits of the BK(Ca) channel may play a critical role in the modulation of channel activity. Therefore, this interface represents a potential therapeutic target site for the regulation of K(+) channels.
[show abstract][hide abstract] ABSTRACT: Quinolinate phosphoribosyltransferase (QPRTase) is a key NAD-biosynthetic enzyme which catalyzes the transfer of quinolinic acid to 5-phosphoribosyl-1-pyrophosphate, yielding nicotinic acid mononucleotide. Homo sapiens QPRTase (Hs-QPRTase) appeared as a hexamer during purification and the protein was crystallized. Diffraction data were collected and processed at 2.8 Å resolution. Native Hs-QPRTase crystals belonged to space group P2(1), with unit-cell parameters a=76.2, b=137.1, c=92.7 Å, β=103.8°. Assuming the presence of six molecules in the asymmetric unit, the calculated Matthews coefficient is 2.46 Å3 Da(-1), which corresponds to a solvent content of 49.9%.
Acta Crystallographica Section F Structural Biology and Crystallization Communications 01/2011; 67(Pt 1):38-40. · 0.55 Impact Factor
[show abstract][hide abstract] ABSTRACT: NAmPRTase (PBEF/Visfatin) plays a pivotal role in the salvage pathway of NAD(+) biosynthesis. NAmPRTase has been an attractive target for anti-cancer agents that induce apoptosis of tumor cells via a declining plasma NAD(+) level. In this report, a series of structural analogs of FK866 (1), a known NAmPRTase inhibitor, was synthesized and tested for inhibitory activities against the proliferation of cancer cells and human NAmPRTase. Among them, compound 7 showed similar anti-cancer and enzyme inhibitory activities to compound 1. Further investigation of compound 7 with X-ray analysis revealed a co-crystal structure in complex with human NAmPRTase, suggesting that Asp219 in the active site of the enzyme could contribute to an additional interaction with the pyrrole nitrogen of compound 7.
European journal of medicinal chemistry 01/2011; 46(4):1153-64. · 3.27 Impact Factor
[show abstract][hide abstract] ABSTRACT: Ryanodine receptors (RyRs) are intracellular Ca(2+) release channels (CRCs) that play a pivotal role in cellular Ca(2+) signaling. In striated muscles, RyR-mediated Ca(2+) release from the sarcoplasmic reticulum (SR) induces elevation of cytosolic Ca(2+) concentration and subsequent muscle contraction. Evidence from various sources suggests that RyRs in homo-tetrameric conformation form a large conductance Ca(2+) permeable channel in the central pore and large cytoplasmic domains. RyRs form a large assembly with various cytosolic and luminal proteins. A number of papers have been published concerning the functions of RyRs and the regulation of the associated proteins, but the three dimensional (3D) structure of the assembly has not been addressed in detail. In this paper, we have attempted to establish a 3D-map for the assembly of RyRs by considering published cryo-EM data, available X-ray crystallographic information and molecular modeling methods.
Progress in Biophysics and Molecular Biology 10/2010; 105(3):145-61. · 2.91 Impact Factor
[show abstract][hide abstract] ABSTRACT: In Gram-negative bacteria, proper placement of the FtsZ ring, mediated by nucleoid occlusion and the activities of the dynamic oscillating Min proteins MinC, MinD and MinE, is required for correct positioning of the cell division septum. MinE is a topological specificity factor that counters the activity of MinCD division inhibitor at the mid-cell division site. Its structure consists of an anti-MinCD domain and a topology specificity domain (TSD). Previous NMR analysis of truncated Escherichia coli MinE showed that the TSD domain contains a long alpha-helix and two anti-parallel beta-strands, which mediate formation of a homodimeric alpha/beta structure. Here we report the crystal structure of full-length Helicobacter pylori MinE and redefine its TSD based on that structure. The N-terminal region of the TSD (residues 19-26), previously defined as part of the anti-MinCD domain, forms a beta-strand (betaA) and participates in TSD folding. In addition, H. pylori MinE forms a dimer through the interaction of anti-parallel betaA-strands. Moreover, we observed serial dimer-dimer interactions within the crystal packing, resulting in the formation of a multimeric structure. We therefore redefine the functional domain of MinE and propose that a multimeric filamentous structure is formed through anti-parallel beta-strand interactions.
[show abstract][hide abstract] ABSTRACT: Oxidative stress is imperative for its morbidity towards diabetic complications, where abnormal metabolic milieu as a result of hyperglycemia, leads to the onset of several complications. A biological antioxidant capable of inhibiting oxidative stress mediated diabetic progressions; during hyperglycemia is still the need of the era. The current study was performed to study the effect of biologically synthesized gold nanoparticles (AuNPs) to control the hyperglycemic conditions in streptozotocin induced diabetic mice.
The profound control of AuNPs over the anti oxidant enzymes such as GSH, SOD, Catalase and GPx in diabetic mice to normal, by inhibition of lipid peroxidation and ROS generation during hyperglycemia evidence their anti-oxidant effect during hyperglycemia. The AuNPs exhibited an insistent control over the blood glucose level, lipids and serum biochemical profiles in diabetic mice near to the control mice provokes their effective role in controlling and increasing the organ functions for better utilization of blood glucose. Histopathological and hematological studies revealed the non-toxic and protective effect of the gold nanoparticles over the vital organs when administered at dosage of 2.5 mg/kilogram.body.weight/day. ICP-MS analysis revealed the biodistribution of gold nanoparticles in the vital organs showing accumulation of AuNPs in the spleen comparatively greater than other organs.
The results obtained disclose the effectual role of AuNPs as an anti-oxidative agent, by inhibiting the formation of ROS, scavenging free radicals; thus increasing the anti-oxidant defense enzymes and creating a sustained control over hyperglycemic conditions which consequently evoke the potential of AuNPs as an economic therapeutic remedy in diabetic treatments and its complications.
Journal of Nanobiotechnology 01/2010; 8:16. · 5.09 Impact Factor