Rajan Sankaranarayanan

Centre for Cellular and Molecular Biology, Bhaganagar, Telangana, India

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Publications (84)352.95 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Proofreading modules of aminoacyl-tRNA synthetases are responsible for enforcing a high fidelity during translation of the genetic code. They use strategically positioned side chains for specifically targeting incorrect aminoacyl-tRNAs. Here, we show that a unique proofreading module possessing a D-aminoacyl-tRNA deacylase fold does not use side chains for imparting specificity or for catalysis, the two hallmark activities of enzymes. We show, using three distinct archaea, that a side-chain-stripped recognition site is fully capable of solving a subtle discrimination problem. While biochemical probing establishes that RNA plays the catalytic role, mechanistic insights from multiple high-resolution snapshots reveal that differential remodelling of the catalytic core at the RNA-peptide interface provides the determinants for correct proofreading activity. The functional crosstalk between RNA and protein elucidated here suggests how primordial enzyme functions could have emerged on RNA-peptide scaffolds before recruitment of specific side chains.
    Nature Communications 01/2015; 6:7552. DOI:10.1038/ncomms8552 · 10.74 Impact Factor
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    ABSTRACT: Xanthomonas oryzae pv. oryzae, the bacterial blight pathogen of rice, secretes a number of effectors through a type 3 secretion system. One of these effectors, called XopQ, is required for virulence and suppression of rice innate immune responses induced by the plant cell wall degrading enzyme LipA (lipase/esterase A). Bioinformatic analysis suggested that XopQ is homologous to inosine-uridine nucleoside hydrolases (NH). A structural model of XopQ with the protozoan Crithidia fasciculata purine NH suggested that D116 and Y279 are potential active site residues. X. oryzae pv. oryzae xopQ mutants (xopQ-/pHM1::xopQD116A and xopQ-/pHM1::xopQY279A) show reduced virulence on rice as compared to xopQ-/pHM1::xopQ. The two predicted XopQ active site mutants (xopQ-/pHM1::xopQD116A and xopQ-/pHM1::xopQY279A) exhibit reduced hypersensitive response (HR) on Nicotiana benthamiana, a non-host. However, Arabidopsis lines expressing either xopQ or xopQY279A are equally proficient at suppression of LipA induced callose deposition. Purified XopQ does not show NH activity on standard nucleoside substrates but exhibits ribose hydrolase activity on the nucleoside substrate analogue 4-nitrophenyl β-D-ribofuranoside. The D116A and Y279A mutations cause a reduction in biochemical activity. These results indicate that mutations in the predicted active site of XopQ affect virulence and induction of HR but do not affect suppression of innate immunity.
    Molecular Plant-Microbe Interactions 10/2014; 28(2). DOI:10.1094/MPMI-09-14-0288-R · 4.46 Impact Factor
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    ABSTRACT: Substrate binding to enzymes often follows a precise order where catalysis is accomplished through programmed conformational changes. Short-chain dehydrogenase/reductase (SDR) enzymes follow sequential order ‘bi–bi’ reaction kinetics. The mechanistic study of a SDR homolog, reductase (R) domain, from multifunctional enzymes, e.g. Nonribosomal Peptide Synthetases (NRPSs) and Polyketide Synthases (PKSs) has revealed that it reductively releases 4′-phosphopantetheinyl arm-tethered peptidyl product. We report that the R-domains of NRPSs from Mycobacterium tuberculosis (RNRP) and Mycobacterium smegmatis (RGPL) do not strictly adhere to the obligatory mode of catalysis performed by SDRs, but instead can carry out reductive catalysis of substrate following random bi–bi reaction mechanism as deciphered by NMR and SAXS studies. The crucial conformational change associated with NADPH binding necessary to achieve catalytically competent conformation is also delineated by SAXS studies. Using ITC, we have demonstrated that mutation of catalytic tyrosine to phenylalanine in R-domains results in 3–4-fold decrease in affinity for NADPH and attribute this phenomenon to loss of the noncovalent cation–π interactions present between the tyrosine and nicotinamide ring. We propose that the adaptation to an alternative theme of bi–bi catalytic mechanism enables the R-domains to process the substrates transferred by upstream domains and maintain assembly-line enzymology.
    Journal of Structural Biology 09/2014; 187(3). DOI:10.1016/j.jsb.2014.07.008 · 3.37 Impact Factor
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    ABSTRACT: eLife digest Amino acids are ‘chiral’ molecules that come in two different forms, called D and L, which are mirror images of each other, similar to how our left and right hands are mirror images of each other. However, only one of these forms is used to make proteins: the more abundant L-amino acids are linked together to make proteins, whereas the scarcer D-amino acids are not. This ‘homochirality’ is common to all life on Earth. The molecular machinery inside cells that manufactures proteins involves many enzymes that carry out different tasks. Among these is an enzyme called DTD (short for D-aminoacyl-tRNA deacylase), which prevents D-amino acids being incorporated into proteins. To do this, DTD must be able to recognise and remove the D forms of many different amino acids before they are taken to the growing protein by transfer RNA molecules. However, the details of this process are not fully understood. To investigate this mechanism, Ahmad et al. made crystals of the DTD enzyme in complex with a molecule that mimics a D-amino acid attached to a transfer RNA molecule. By studying this structure at a high resolution, Ahmad et al. were able to identify how the active site of DTD can specifically accommodate the ‘chiral centre’ of a complex made of a D-amino acid and a transfer RNA molecule. DTD is able to recognize D-amino acids because of a critical dipeptide that is inserted from one subunit of the DTD into the active site of another subunit of the enzyme. The effect of this dipeptide is to generate a binding pocket that is a perfect fit for the chiral centre of a complex that contains a D-amino acid and a transfer RNA molecule. Moreover, this pocket specifically excludes complexes that contain an L-amino acid. The crucial parts of DTD that form the binding pocket are highly conserved—that is, they are the same in a wide variety of organisms, from bacteria to mammals. This conservation suggests that DTD is crucial for ensuring homochirality throughout all forms of life. Intriguingly, DTD is particularly highly expressed in neurons which are abundant in D-amino acids: this indicates that the DTD enzyme has an important physiological role, which will certainly be the focus of future work. DOI: http://dx.doi.org/10.7554/eLife.01519.002
    eLife Sciences 12/2013; 2:e01519. DOI:10.7554/eLife.01519 · 8.52 Impact Factor
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    ABSTRACT: The proofreading function of aminoacyl-tRNA synthetases is crucial in maintaining the fidelity of protein synthesis. Most archaeal threonyl-tRNA synthetases (ThrRSs) possess a unique proofreading domain unrelated to their eukaryotic/bacterial counterpart. The crystal structure of this domain from the archaeon Pyrococcus abysii in complex with its cognate and noncognate substrate analogues had given insights into its catalytic and discriminatory mechanisms. To probe further into the mechanistic and evolutionary aspects of this domain, work has been extended to another archaeon Aeropyrum pernix. The organism possesses two proteins corresponding to threonyl-tRNA synthetase, i.e. ThrRS1 and ThrRS2, encoded by two different genes, thrS1 and thrS2, respectively. ThrRS1 is responsible for aminoacylation and ThrRS2 for proofreading activity. Here the purification, crystallization and preliminary X-ray crystallographic investigation of the N-terminal proofreading domain of ThrRS2 from A. pernix is reported. The crystals belong to either the P4(1)2(1)2 or P4(3)2(1)2 space group and consist of one monomer per asymmetric unit.
    Acta Crystallographica Section F Structural Biology and Crystallization Communications 11/2012; 68(Pt 11):1390-1393. DOI:10.1107/S1744309112042066 · 0.57 Impact Factor
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    ABSTRACT: The folding and unfolding of structurally similar proteins belonging to a family have long been a focus of investigation of the structure-(un)folding relationship. Such studies are yet to reach a consensus about whether structurally similar domains follow common or different unfolding pathways. Members of the βγ-crystallin superfamily, which consists of structurally similar proteins with limited sequence similarity from diverse life forms spanning microbes to mammals, form an appropriate model system for exploring this relationship further. We selected a new member, Crybg3_D3, the third βγ-crystallin domain of non-lens vertebrate protein Crybg3 from mouse brain. The crystal structure determined at 1.86 Å demonstrates that the βγ-crystallin domain of Crybg3 resembles more closely the lens βγ-crystallins than the microbial crystallins do. However, interestingly, this structural cousin follows a quite distinct (un)folding pathway via formation of an intermediate state. The intermediate species is in a nativelike conformation with variation in flexibility and tends to form insoluble aggregates. The individual domains of lens βγ-crystallins (and microbial homologues) do not follow such an unfolding pattern. Thus, even the closest members of a subfamily within a superfamily do not necessarily follow similar unfolding paths, suggesting the divergence acquired by these domains, which could be observed only by unfolding. Additionally, this study provides insights into the modifications that this domain has undergone during its recruitment into the non-lens tissues in vertebrates.
    Biochemistry 10/2012; 51(43). DOI:10.1021/bi300844u · 3.19 Impact Factor
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    ABSTRACT: The bacterial pathogen Xanthomonas oryzae pv. oryzae causes bacterial leaf blight, a serious disease of rice. The secreted exoglucanase CbsA is an important virulence factor of this pathogen. It belongs to the glycosyl hydrolase 6 family of proteins based on the carbohydrate-active enzyme (CAZY) classification. In this study, CbsA has been overexpressed, purified and crystallized. The crystal diffracted to a resolution of 1.86 Å and belonged to space group P2(1)2(1)2(1). It contained one monomer per asymmetric unit, with a solvent content of 45.8%.
    Acta Crystallographica Section F Structural Biology and Crystallization Communications 10/2012; 68(Pt 10):1191-4. DOI:10.1107/S1744309112034197 · 0.57 Impact Factor
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    ABSTRACT: In mycobacteria, polyketide synthases and nonribosomal peptide synthetases (NRPSs) produce complex lipidic metabolites by using a thio-template mechanism of catalysis. In this study, we demonstrate that off-loading reductase (R) domain of mycobacterial NRPSs performs two consecutive [2 + 2]e(-) reductions to release thioester-bound lipopeptides as corresponding alcohols, using a nonprocessive mechanism of catalysis. The first crystal structure of an R domain from Mycobacterium tuberculosis NRPS provides strong support to this mechanistic model and suggests that the displacement of intermediate would be required for cofactor recycling. We show that 4e(-) reductases produce alcohols through a committed aldehyde intermediate, and the reduction of this intermediate is at least 10 times more efficient than the thioester-substrate. Structural and biochemical studies also provide evidence for the conformational changes associated with the reductive cycle. Further, we show that the large substrate-binding pocket with a hydrophobic platform accounts for the remarkable substrate promiscuity of these domains. Our studies present an elegant example of the recruitment of a canonical short-chain dehydrogenase/reductase family member as an off-loading domain in the context of assembly-line enzymology.
    Proceedings of the National Academy of Sciences 03/2012; 109(15):5681-6. DOI:10.1073/pnas.1118680109 · 9.81 Impact Factor
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    ABSTRACT: Edited by J. E. Ladbury Keywords: βγ-crystallin; calcium binding; calcium-binding motif; N/D-N/D-X-X-S/T-S motif Numerous proteins belonging to the recently expanded βγ-crystallin superfamily bind Ca 2+ at the double-clamp N/D-N/D-X 1 -X 2 -S/T-S motif. However, there have been no attempts to understand the intricacies involving Ca 2+ binding, such as the determinants of Ca 2+ -binding affinity and their contributions to gain in stability. This work is an in-depth analysis of understanding the modes and determinants of Ca 2+ binding to βγ-crystallin motifs. We have performed extensive naturally occurring substitutions from related proteins on the βγ-crystallin domains of flavollin, a low-affinity Ca 2+ -binding protein, and clostrillin, a moderate-affinity protein. We monitored the consequences of these modifications on Ca 2 + binding by isothermal titration calorimetry, thermal stability and confor-mational and crystal structure analyses. We demonstrate that Ca 2 + binding to the two sites of a βγ-domain is interdependent and that the presence of Arg at the fifth position disables a site. A change from Thr to Ser, or vice versa, influences Ca 2+ -binding affinity, highlighting the basis of diversity found in these domains. A subtle change in the first site has a greater influence on Ca 2 + binding than a similar alteration in the second site. Thus, the second site is more variable in nature. Replacing an acidic or hydrophobic residue in a binding site alters the Ca 2+ -binding properties drastically. While it appears from their binding site sequence that these domains have evolved randomly, our examination illustrates the subtlety in the design of these modules. Decoding such design schemes would aid in our understanding of the functional themes underlying differential Ca 2 + binding and in predicting these in emerging sequence information.
    Journal of Molecular Biology 01/2012; · 3.96 Impact Factor
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    ABSTRACT: Activation of fatty acids as acyl-adenylates by fatty acyl-AMP ligases (FAALs) in Mycobacterium tuberculosis is a variant of a classical theme that involves formation of acyl-CoA (coenzyme A) by fatty acyl-CoA ligases (FACLs). Here, we show that FAALs and FACLs possess similar structural fold and substrate specificity determinants, and the key difference is the absence of a unique insertion sequence in FACL13 structure. A systematic analysis shows a conserved hydrophobic anchorage of the insertion motif across several FAALs. Strikingly, mutagenesis of two phenylalanine residues, which are part of the anchorage, to alanine converts FAAL32 to FACL32. This insertion-based in silico analysis suggests the presence of FAAL homologues in several other non-mycobacterial genomes including eukaryotes. The work presented here establishes an elegant mechanism wherein an insertion sequence drives the functional divergence of FAALs from canonical FACLs.
    Journal of Molecular Biology 12/2011; 416(2):221-38. DOI:10.1016/j.jmb.2011.12.031 · 3.96 Impact Factor
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    ABSTRACT: Leishmania META1 has for long been a candidate molecule for involvement in virulence: META1 transcript and protein are up-regulated in metacyclic Leishmania. Yet, how META1 contributes to virulence remains unclear. We sought insights into the possible functions of META1 by studying its evolutionary origins. Using multiple criteria including sequence similarity, nucleotide composition, phylogenetic analysis and selection pressure on gene sequence, we present evidence that META1 originated in trypanosomatids as a result of a lateral gene transfer of a bacterial heat-inducible protein, HslJ. Furthermore, within the Leishmania genome, META1 sequence is under negative selection pressure against change/substitution. Using homology modeling of Leishmania META1 based on solved NMR structure of HslJ, we show that META1 and HslJ share a similar structural fold. The best hit for other proteins with similar fold is MxiM, a protein involved in the type III secretion system in Shigella. The striking structural similarity shared by META1, HslJ and MxiM suggests a possibility of shared functions. Upon structural superposition with MxiM, we have observed a putative hydrophobic cavity in META1. Mutagenesis of select hydrophobic residues in this cavity affects the secretion of the secreted acid phosphatase (SAP), indicating META1's involvement in secretory processes in Leishmania. Overall, this work uses an evolutionary biology approach, 3D-modeling and site-directed mutagenesis to arrive at new insights into functions of Leishmania META1.
    BMC Evolutionary Biology 11/2011; 11:334. DOI:10.1186/1471-2148-11-334 · 3.41 Impact Factor
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    ABSTRACT: Numerous proteins belonging to the recently expanded βγ-crystallin superfamily bind Ca(2+) at the double-clamp N/D-N/D-X(1)-X(2)-S/T-S motif. However, there have been no attempts to understand the intricacies involving Ca(2+) binding, such as the determinants of Ca(2+)-binding affinity and their contributions to gain in stability. This work is an in-depth analysis of understanding the modes and determinants of Ca(2+) binding to βγ-crystallin motifs. We have performed extensive naturally occurring substitutions from related proteins on the βγ-crystallin domains of flavollin, a low-affinity Ca(2+)-binding protein, and clostrillin, a moderate-affinity protein. We monitored the consequences of these modifications on Ca(2)(+) binding by isothermal titration calorimetry, thermal stability and conformational and crystal structure analyses. We demonstrate that Ca(2)(+) binding to the two sites of a βγ-domain is interdependent and that the presence of Arg at the fifth position disables a site. A change from Thr to Ser, or vice versa, influences Ca(2+)-binding affinity, highlighting the basis of diversity found in these domains. A subtle change in the first site has a greater influence on Ca(2)(+) binding than a similar alteration in the second site. Thus, the second site is more variable in nature. Replacing an acidic or hydrophobic residue in a binding site alters the Ca(2+)-binding properties drastically. While it appears from their binding site sequence that these domains have evolved randomly, our examination illustrates the subtlety in the design of these modules. Decoding such design schemes would aid in our understanding of the functional themes underlying differential Ca(2)(+) binding and in predicting these in emerging sequence information.
    Journal of Molecular Biology 10/2011; 415(1):75-91. DOI:10.1016/j.jmb.2011.10.037 · 3.96 Impact Factor
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    ABSTRACT: Rational and in vitro evolutionary approaches to improve either protein stability or aggregation resistance were successful, but empirical rules for simultaneous improvement of both stability and aggregation resistance under denaturing conditions are still to be ascertained. We have created a robust variant of a lipase from Bacillus subtilis named "6B" using multiple rounds of in vitro evolution. T(m) and optimum activity temperature of 6B is 78 °C and 65 °C, respectively, which is ~22 °C and 30 °C higher than that of wild-type lipase. Most significantly, 6B does not aggregate upon heating. Physical basis of remarkable thermostability and non-aggregating behavior of 6B was explored using X-ray crystallography, NMR and differential scanning calorimetry. Our structural investigations highlight the importance of tightening of mobile regions of the molecule such as loops and helix termini to attain higher thermostability. Accordingly, NMR studies suggest a very rigid structure of 6B lipase. Further investigation suggested that reduction/perturbation of the large hydrophobic patches present in the wild-type protein structure, decreased propensity of amino acid sequence for aggregation and absence of aggregation-prone intermediate during thermal unfolding of 6B can account for its resistance to aggregation. Overall, our study suggest that better anchoring of the loops with the rest of the protein molecule through mutations particularly on the sites that perturb/disturb the exposed hydrophobic patches can simultaneously increase protein stability and aggregation resistance.
    Journal of Molecular Biology 09/2011; 413(3):726-41. DOI:10.1016/j.jmb.2011.09.002 · 3.96 Impact Factor
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    ABSTRACT: NGP-1(GNL-2) is a putative GTPase, overexpressed in breast carcinoma and localized in the nucleolus. NGP-1 belongs to the MMR1-HSR1 family of large GTPases that are emerging as crucial coordinators of signaling cascades in different cellular compartments. The members of this family share very closely related G-domains, but the signals and pathways regulating their subcellular localization and their functional relevance remain unknown. To improve our understanding of the nuclear transport mechanism of NGP-1, we have identified two nucleolar localization signals (NoLS) that are independently shown to translocate NGP-1 as well the heterologous protein to the nucleolus. Site-specific mutagenesis and immunofluorescence studies suggest that the tandem repeats of positively charged amino acids are critical for NGP-1 NoLS function. Interestingly, amino-terminal (NGP-1(1-100)) and carboxyl-terminal (NGP-1(661-731)) signals independently interact with receptors importin-β and importin-α, respectively. This investigation, for the first time, provides evidence that the interaction of importin-α with C-terminal NoLS (NGP-1(661-731)) was able to target the heterologous protein to the nucleolar compartment. Structural modeling analysis and alanine scanning mutagenesis of conserved G-domains suggest that G4 and G5 motifs are critical for GTP binding of NGP-1 and further show that the nucleolar localization of NGP-1 is regulated by a GTP gating-mediated mechanism. In addition, our data suggest that an ongoing transcription is essential for efficient localization of NGP-1 to the nucleolus. We have observed a high level of NGP-1 expression in the mitogen-activated primary human peripheral blood mononuclear cells (hPBMC) as well as in human fetal brain-derived neural precursor cells (hNPCs) in comparison to cells undergoing differentiation. Overall, the results suggest that multiple mechanisms are involved in the localization of NGP-1 to the nucleolus for the regulation of nucleolar function in cell growth and proliferation.
    Biochemistry 05/2011; 50(21):4521-36. DOI:10.1021/bi200425b · 3.19 Impact Factor
  • Debasisa Mohanty, Rajan Sankaranarayanan, Rajesh S Gokhale
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    ABSTRACT: The cell envelope of Mycobacterium tuberculosis (Mtb) possesses a repertoire of unusual lipids that are believed to play an important role in pathogenesis. In this review, we specifically focus on computational, biochemical and structural studies in lipid biosynthesis that have established functional role of polyketide synthases (PKSs) and fatty acyl-AMP ligases (FAALs). Mechanistic and structural studies with FAALs suggest that this group of proteins may have evolved from omnipresent fatty acyl-CoA ligases (FACLs). FAALs activate fatty acids as acyl-adenylates and transfer them on to the PKSs which then produce unusual acyl chains that are the components of mycobacterial lipids. FAALs are a newly discovered family of enzymes; whereas involvement of PKSs in lipid metabolism was not known prior to their discovery in Mtb. Since Mtb genome contains multiple homologs of FAALs and PKSs and owing to the conserved reaction mechanism and overlapping substrate specificity; there is tempting opportunity to develop 'systemic drugs' against these enzymes as anti-tuberculosis agents.
    Tuberculosis (Edinburgh, Scotland) 05/2011; 91(5):448-55. DOI:10.1016/j.tube.2011.04.006 · 3.50 Impact Factor
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    ABSTRACT: Among the many different objectives of large scale structural genomics projects are expanding the protein fold space, enhancing understanding of a model or disease-related organism, and providing foundations for structure-based drug discovery. Systematic analysis of protein structures of Mycobacterium tuberculosis has been ongoing towards meeting some of these objectives. Indian participation in these efforts has been enthusiastic and substantial. The proteins of M. tuberculosis chosen for structural analysis by the Indian groups span almost all the functional categories. The structures determined by the Indian groups have led to significant improvement in the biochemical knowledge on these proteins and consequently have started providing useful insights into the biology of M. tuberculosis. Moreover, these structures form starting points for inhibitor design studies, early results of which are encouraging. The progress made by Indian structural biologists in determining structures of M. tuberculosis proteins is highlighted in this review.
    Tuberculosis (Edinburgh, Scotland) 04/2011; 91(5):456-68. DOI:10.1016/j.tube.2011.03.004 · 3.50 Impact Factor
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    ABSTRACT: Editing/proofreading by aminoacyl-tRNA synthetases is an important quality control step in the accurate translation of the genetic code that removes noncognate amino acids attached to tRNA. Defects in the process of editing result in disease conditions including neurodegeneration. While proofreading, the cognate amino acids larger by a methyl group are generally thought to be sterically rejected by the editing modules as envisaged by the "Double-Sieve Model." Strikingly using solution based direct binding studies, NMR-heteronuclear single quantum coherence (HSQC) and isothermal titration calorimetry experiments, with an editing domain of threonyl-tRNA synthetase, we show that the cognate substrate can gain access and bind to the editing pocket. High-resolution crystal structural analyses reveal that functional positioning of substrates rather than steric exclusion is the key for the mechanism of discrimination. A strategically positioned "catalytic water" molecule is excluded to avoid hydrolysis of the cognate substrate using a "RNA mediated substrate-assisted catalysis mechanism" at the editing site. The mechanistic proof of the critical role of RNA in proofreading activity is a completely unique solution to the problem of cognate-noncognate selection mechanism.
    Proceedings of the National Academy of Sciences 12/2010; 107(51):22117-21. DOI:10.1073/pnas.1014299107 · 9.81 Impact Factor
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    ABSTRACT: ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.
    ChemInform 07/2010; 31(30). DOI:10.1002/chin.200030031
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    ABSTRACT: Polycomb group (PcG) and trithorax group (trxG) proteins are chromatin-mediated regulators of a number of developmentally important genes including the homeotic genes. In Drosophila melanogaster, one of the trxG members, Trithorax like (Trl), encodes the essential multifunctional DNA binding protein called GAGA factor (GAF). While most of the PcG and trxG genes are conserved from flies to humans, a Trl-GAF homologue has been conspicuously missing in vertebrates. Here, we report the first identification of c-Krox/Th-POK as the vertebrate homologue of GAF on the basis of sequence similarity and comparative structural analysis. The in silico structural analysis of the zinc finger region showed preferential interaction of vertebrate GAF with GAGA sites similar to that of fly GAF. We also show by cross-immunoreactivity studies that both fly and vertebrate GAFs are highly conserved and share a high degree of structural similarity. Electrophoretic mobility shift assays show that vertebrate GAF binds to GAGA sites in vitro. Finally, in vivo studies by chromatin immunoprecipitation confirmed that vertebrate GAF binds to GAGA-rich DNA sequences present in hox clusters. Identification of vertebrate GAF and the presence of its target sites at various developmentally regulated loci, including hox complexes, highlight the evolutionarily conserved components involved in developmental mechanisms across the evolutionary lineage and answer a long-standing question of the presence of vertebrate GAF.
    Journal of Molecular Biology 07/2010; 400(3):434-47. DOI:10.1016/j.jmb.2010.05.010 · 3.96 Impact Factor
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    ABSTRACT: Proteins belonging to the lipocalin superfamily are usually secretory proteins of molecular mass approximately 20 kDa with a hydrophobic pocket for the binding and transport of diverse small ligands. Various lipocalins have been associated with many biological processes, e.g. immunomodulation, odorant transport, pheromonal activity, retinoid transport, cancer-cell interactions etc. However, the exact functions of many lipocalins and the ligands bound by them are unclear. Previously, the cDNA of a 20 kDa lipocalin (FLP) which is female-specifically expressed in the lacrimal glands of Syrian (golden) hamsters and secreted in the tears of females has been identified and cloned. His-tagged recombinant FLP (rFLP) has now been cloned, overexpressed in Escherichia coli as a soluble protein and purified to homogeneity using Ni-affinity followed by size-exclusion chromatography. Purified rFLP was crystallized using the sitting-drop vapour-diffusion method. The crystals tested belonged to space group P2(1)2(1)2(1) and diffracted to beyond 1.86 A resolution. Solvent-content analysis indicated the presence of one monomer in the asymmetric unit.
    Acta Crystallographica Section F Structural Biology and Crystallization Communications 05/2010; 66(Pt 5):509-12. DOI:10.1107/S1744309110008237 · 0.57 Impact Factor

Publication Stats

2k Citations
352.95 Total Impact Points

Institutions

  • 2004–2015
    • Centre for Cellular and Molecular Biology
      Bhaganagar, Telangana, India
  • 2004–2011
    • Molecular and Cellular Biology Program
      • Department of Molecular and Cellular Biology
      Seattle, Washington, United States
  • 1999–2010
    • University of Madras
      • Department of Crystallography and Biophysics
      Chennai, Tamil Nādu, India
    • French National Centre for Scientific Research
      • Institute for Molecular and Cellular Biology (IBMC)
      Lutetia Parisorum, Île-de-France, France
  • 2009
    • University of Alberta
      • Department of Biochemistry
      Edmonton, Alberta, Canada
  • 2008
    • National Institutes of Health
      Maryland, United States
  • 2005
    • IIT Kharagpur
      • Department of Biotechnology
      Kharagpur, Bengal, India
  • 2000–2003
    • Institut de Génétique et de Biologie Moléculaire et Cellulaire
      • Department of Functional Genomics and Cancer
      Strasburg, Alsace, France
  • 2001
    • French Institute of Health and Medical Research
      Lutetia Parisorum, Île-de-France, France