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130
Elena Cressina, Liuhong Chen, Michael Moulin, Finian J Leeper, Chris Abell and Alison G Smith, “Identification of novel ligands for thiamine pyrophosphate (TPP) riboswitches” Biochem. Soc. Trans., 2011, 39 (2), 652-7. Full Text.

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.

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131
Henning Stöckmann, Andre A. Neves, Henry A. Day, Shaun Stairs, Kevin M. Brindle and and F. J. Leeper, "(E,E)-1,5-cyclooctadiene: A small and fast click-chemistry multitalent" Chem. Comm., 2011, 47 (25), 7203-7205. Full Text.

Two in one – We show here that the highly strained trans,trans-diolefin (E,E)-1,5-cyclooctadiene can perform efficiently two different click reactions at fast reaction rates. It is capable of first undergoing [3+2] cycloadditions with 1,3-dipoles at a reaction rate comparable to that of strained cyclooctynes. The resulting cycloadduct can then perform a much faster inverse-electrondemand Diels-Alder reaction with tetrazines, effectively linking an azide to a tetrazine. Thus, (E,E)-1,5-cyclooctadiene could have many applications in chemical biology and polymer chemistry.

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132
Henning Stöckmann, André A. Neves, Shaun Stairs, Kevin M. Brindle and Finian J Leeper, "Exploring isonitrile-based click chemistry for ligation with biomolecules" Org. Biomol. Chem., 2011, 9 (21), 7303-7305. Full Text.

We show here that isonitriles can perform click reactions with tetrazines in aqueous media, making them promising candidates for ligation reactions in chemical biology and polymer chemistry. This is the first time that a [4+1] cycloaddition has been used as a biocompatible ligation reaction.

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133
Suresh R. Chawrai, Neil R. Williamson, Thabo Mahendiran, George P. C. Salmond and Finian J Leeper, "Characterisation of PigC and HapC, the prodigiosin synthetases from Serratia sp. and Hahella chejuensis with potential for biocatalytic production of anticancer agents" Chem. Sci., 2012, 3 (2), 447-454. Full Text.

PigC and HapC catalyse the condensation of 4-methoxy-2,2’-bipyrrole-5-carbaldehyde (MBC) with 2- methyl-3-amylpyrrole (MAP) to give the bright red tripyrrolic prodigiosin, which has potent anti-cancer activity. We have cloned and over-expressed both enzymes and characterised their enzymic activity in vitro using both the natural substrates, MBC and MAP, and analogues of these substrates. Thus a range of prodigiosin analogues have been produced. Both PigC and HapC are membrane-associated enzymes and attempts to fully solubilise them using detergents led to inactivation. The enzymes are ATP-dependent but, unlike the enzymes to which they show the greatest similarity, the by-product is ADP not AMP. Two different slowly interconverting rotamers of prodigiosin exist and the spectral changes with time are consistent with isomerisation of the E,Z (or a) rotamer to the Z,Z (or b) rotameric form.

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134
Finian J Leeper, Martin J. Warren, James M. Kelly and Andrew D. Lawrence
"Biosynthesis of Vitamin B₁₂ "
in Handbook of Porphyrin Science, eds. Karl M. Kadish and Kevin M. Smith, 2012, in press.

Vitamin B12 is required by mammals but it is only made by bacteria, hence it is a vitamin that is mandated in the human diet. Its structure, first solved by Dorothy Hodgkin's group using X-ray crystallography, was for a long time the most complex non-repetitive natural product known. The total syntheses of vitamin B12 by the groups of Woodward and of Eschenmoser were regarded as the pinnacle of achievement in organic synthesis, and elucidation of the biosynthesis of the vitamin was also a tremendous challenge. This chapter gives an account of the current knowledge of the biosynthesis as well as some of the historical background of the field.

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135
James M. Kelly and Finian J Leeper,
"Synthesis of 3,4-fused cycloalkanopyrroles by 1,3-dipolar cycloaddition"
Tetrahedron Lett., 2012, 53 (7), 819-821. Full Text.

The synthesis of a number of 3,4-fused cycloalkanopyrroles bearing substituents on the cycloalkane ring was accomplished by 1,3-dipolar cycloaddition. The yield of the cyclization appeared to depend on the base-sensitivity of the Michael acceptor, but the method is applicable across a broad range of cyclic ?,?-unsaturated ketone esters. Functional group transformations can be undertaken following pyrrole synthesis to increase the diversity of cycloalkanopyrroles accessible by this method. One pyrrole thus made is an diester of a conformational constrained analogue of porphobilinogen, the precursor of the natural tetrapyrroles.

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136
Helen L. Evans, Rozanna L. Slade, Laurence Carroll, Graham Smith, Quang-De Nguyen, Lisa Iddon, Nazilla Kamaly, Henning Stöckmann, Finian J. Leeper, Eric O. Aboagye and Alan C. Spivey,
"Copper-free Click – A Promising Tool for Pre-Targeted PET Imaging"
Chem. Comm., 2012, 48 (7), 991-993. Full Text.

The copper-free click (CFC) reaction has been evaluated for its potential application to in vivo pre-targeting for PET imaging. A promising biodistribution profile is demonstrated when employing [¹⁸F]2-fluoroethylazide ([¹⁸F]1) and optimisation of the CFC reaction with a series of cyclooctynes shows that reactions proceed efficiently with tantalizing opportunities for application-specific tuning.

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137
Liuhong Chen, Elena Cressina, Neil Dixon, Karl Erixon, Kwasi Agyei-Owusu, Jason Micklefield, Alison G. Smith, Chris Abell and Finian J. Leeper,
"Probing riboswitch–ligand interactions using thiamine pyrrophosphate analogues"
Org. Biomol. Chem., 2012, (), in press. Full Text.

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 ?-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.

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