Elena Cressina

University of Cambridge, Cambridge, ENG, United Kingdom

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Publications (6)25.07 Total impact

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    ABSTRACT: The Escherichia coli thiM riboswitch forms specific contacts with its natural ligand, thiamine pyrophosphate (TPP or thiamine diphosphate), allowing it to generate not only nanomolar binding affinity, but also a high degree of discrimination against similar small molecules. A range of synthetic TPP analogues have been used to probe each of the riboswitch-ligand interactions. The results show that the pyrimidine-sensing helix of thiM is exquisitely tuned to select for TPP by recognising the H-bonding donor and acceptors around its aminopyrimidine ring and also by forming π-stacking interactions that may be sensitive to the electronics of the ring. The central thiazolium ring of TPP appears to be more important for ligand recognition than previously thought. It may contribute to binding via long-range electrostatic interactions and/or by exerting an electron withdrawing effect on the pyrimidine ring, allowing its presence to be sensed indirectly and thereby allowing discrimination between thiamine (and its phosphate esters) and other aminopyrimidines found in vivo. The pyrophosphate moiety is essential for submicromolar binding affinity, but unexpectedly, it does not appear to be strictly necessary for modulation of gene expression.
    Organic & Biomolecular Chemistry 04/2012; 10(30):5924-31. · 3.57 Impact Factor
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    ABSTRACT: Riboswitches are regions of mRNA to which a metabolite binds in the absence of proteins, resoulting in alteration of transcription, translation or splicing. The most widespread forms of riboswitches are those responsive to TPP (thiamine pyrophosphate) the active form of vitamin B1, thiamine. TPP-riboswitches have been found in all bacterial genomes examined, and are the only ones found in eukaryotes. In each case, the riboswitch appears to regulate the expression of a gene involved in synthesis or uptake of the vitamin. Riboswitches offer an attractive target for chemical intervention, and identification of novel ligands would allow a detailed study on structure-activity relationships, as well as potential leads for the development of antimicrobial compounds. To this end, we have developed a medium-throughput methodology for screening libraries of small molecules using biophysical methods.
    Biochemical Society Transactions 04/2011; 39(2):652-7. · 2.59 Impact Factor
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    ABSTRACT: Riboswitches are regions of mRNA that bind selectively certain metabolites, thereby regulating gene expression. We have developed a method, which uses a combination of biophysical techniques (equilibrium dialysis, waterLOGSY and T2 relaxation-edited NMR spectroscopy and isothermal titration calorimetry) that allows screening and identification of novel ligands for riboswitches. By this method a library of 1300 fragments was screened on the E. coli thiamine pyrophosphate (TPP) riboswitchthiM, resulting in the identification of 17 hits. The compounds showed KD values from 22 to 670 μM, with four compounds having KD values <100 μM. The fragments are structurally diverse and their ligand efficiency indicates good prospects for structure-guided elaboration. In addition, a riboswitch functional assay based on in vitro transcriptiontranslation (IVTT) of the reporter geneluciferase was developed. This assay system is an alternative to in vivo assays and constitutes a valuable tool to determine the effect of small molecules on riboswitch-regulated gene expression.
    Chemical Science 12/2010; 2(1):157-165. · 8.31 Impact Factor
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    ABSTRACT: Riboswitches are regions of mRNA that directly bind metabolites, leading to alteration of gene expression. We have developed fragment-based methods to screen for compounds that bind the Escherichia coli thiM riboswitch. Using complementary biophysical techniques we have identified several ligands with K(D) <100 microM. From these there is the potential to develop potent and selective modulators of riboswitch function.
    ACS Chemical Biology 02/2010; 5(4):355-8. · 5.44 Impact Factor
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    ABSTRACT: Ligase MurM catalyses the addition of Ala from alanyl-tRNA(Ala), or Ser from seryl-tRNA(Ser), to lipid intermediate II in peptidoglycan biosynthesis in Streptococcus pneumoniae, and is a determinant of high-level penicillin resistance. Phosphorus-based transition state analogues were designed as inhibitors of the MurM-catalysed reaction. Phosphonamide analogues mimicking the attack of a lysine nucleophile upon Ala-tRNA(Ala) showed no inhibition of MurM, but adenosine 3'-phosphonate analogues showed inhibition of MurM, the most active being a 2'-deoxyadenosine analogue (IC(50) 100 microM). Structure/function studies upon this analogue established that modification of the amino group of the aminoalkylphosphonate resulted in loss of potency, and modification of the adenosine 5'-hydroxyl group with either a t-butyl dimethyl silyl or a carbamate functional group resulted in loss of activity. A library of 48 aryl sulfonamides was also screened against MurM using a radiochemical assay, and two compounds showed sub-millimolar inhibition. These compounds are the first small molecule inhibitors of the Fem ligase family of peptidyltransferases found in Gram-positive bacteria.
    Bioorganic & medicinal chemistry 04/2009; 17(9):3443-55. · 2.82 Impact Factor
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    ABSTRACT: Adenosine and 2'-deoxyadenosine phosphonate transition state analogues act as the first inhibitors for the MurMN/FemABX family of tRNA-dependent ligases implicated in high-level penicillin resistance in gram-positive bacteria.
    Bioorganic & Medicinal Chemistry Letters 09/2007; 17(16):4654-6. · 2.34 Impact Factor