Prof Jason Micklefield

Lecturer in Organic Chemistry, Birkbeck College, University of London (1995 - 1998). NATO Postdoctoral Fellow, University of Washington, Seattle, USA. (1993 - 1994). PhD University of Cambridge, (Clare College) (1993). BSc University of Hull (1989).

The Micklefield research group is engaged in diverse challenges at the chemistry-biology interface, utilising techniques and knowledge from organic chemistry including synthesis through to biochemistry and molecular biology. The main research themes include: biosynthesis and biosynthetic engineering; development and evolution of new biocatalysts; nucleic acid redesign, recognition and template directed synthesis.
 
Biosynthesis and biosynthetic engineering. The Micklefield group is interested in the biosynthesis of nonribosomal peptides. These secondary metabolites include many important therapeutic agents, which are often too complex for effective synthesis. On the other hand, the reprogrammed, engineered biosynthesis of nonribosomal peptide variants, with altered and possibly improved biological activity, is more realistic. Notably the group has elucidated the biosynthetic origins of the calcium dependant antibiotics (CDA) from Streptomyces coelicolor. CDA belongs to the nonribosomal lipopeptide antibiotic family that includes daptomycin, which was introduced into the clinic in 2004 and has become one of the most successful intravenous antibiotics launched in US history [http://www.cubist.com/products/]. Despite this, the occurrence of resistance to daptomycin is widespread in the environment. It is thus essential that new approaches are developed which will enable the production of the next generation lipopeptides that can combat emerging pathogens. To this end the Micklefield group introduced a wide range of methods resulting in the combinatorial biosynthesis of a large number of new lipopeptide antibiotics. Indeed it is possible to make targeted modifications to specific amino acid and fatty acid components of these lipopeptides using the approaches developed. Finally extensive NMR spectroscopy studies have shown that the 3D solution structure of CDA is remarkably similar to that of daptomycin, suggesting a similar mechanism of action. Currently the combined structural and biosynthetic insights are being used to optimise the properties of the next generation lipopeptide antibiotics.

Nucleic acid redesign, recognition and template directed synthesis.A major focus of the Micklefield lab has been the synthesis, conformational analysis and biophysical evaluation of modified nucleic acids and oligonucleotide mimics for therapeutic, diagnostic, bioanalytical and molecular biology applications. For example novel pyrrolidine-amide oligonucleotide mimics (POM) have been introduced, which contain a pyrrolidine-amide backbone that is protonated at physiological pH, rather than the native ribose-phosphodiester structure. POM is therefore cationic and highly soluble in water. NMR and X-ray crystallography have also shown that POM, is conformationally similar to RNA. Moreover biophysical studies (UV, ITC, SPR & CD) reveal that POM binds with extremely high affinity to both native nucleic acids and exhibits remarkable kinetic binding selectivity for RNA over DNA. Most recently the efficient delivery of POM into the nucleus of HeLa cells has been demonstrated. The Micklefield group is also exploring methods, which will allow for the non-enzymatic transcription of abiological polymers (nucleic acid mimics) using dynamic combinatorial chemistry on a DNA template. This approach offers the potential to couple the techniques of molecular biology with organic synthesis to create genetically coded abiological macromolecules with new functions. In addition the template directed (nonenzymatic transcription) chemistry could also be used for high throughput DNA sequencing, towards individual genome sequencing and personalised medicine.

Small molecule modulators of gene expression which offer rapid, temporal and spatial control of gene expression, are important tools in biological and medical sciences. In particular the ability to control differentially the expression of multiple genes simultaneously, using distinct small molecule inhibitors or inducers, would be extremely valuable. Unfortunately very few systems exist with these properties. To this end the Micklefield lab is co-evolving small molecule responsive riboswitches (structured mRNA elements in untranslated regions) that function in prokaryotic and eukaryotic cells and respond in a rapid dose-dependent fashion to a wide range of synthetic "designer" ligands. This could have many profound applications in target validation, gene therapy, biosensors and plant biotechnology.

Biocatalysis.The Micklefield group is interested in the mechanism of enzymes, including, decarboxylase, racemases, epoxidases and mutases and their applications in synthesis. For example methods of directed evolution and genetic selection are used to generate new enzymes with improved properties and altered substrate specificity, that produce valuable homochiral products. Finally the group was the first to demonstrate the significant advantages of pressurised liquid hydrofluorocarbons as a new media for biocatalysis.

Those interested in joining our research group at the PhD or Postdoctoral level should forward their CV and the names of two referees to: Dr Jason Micklefield, MIB, University of Manchester, 131 Princess Street, Manchester - M1 7DN (Tel:  0161 3064509, Email:  jason.micklefield@manchester.ac.uk).

• L. S. Wong, Jenny Thirlway and J. Micklefield, "Site-selective covalent protein immobilization catalysed by a promiscuous phosphopantetheinyl transferase", J. Am. Chem. Soc., 2008, 130, 12456-12464. (PDF)
• S.A.A.Ahmad, L.S.Wong, E. ul-Haq, J.K.Hobbs, G.J.Leggett, J.Micklefield, "Micrometer- and Nanometer-Scale Photopatterning Using 2-Nitrophenylpropyloxycarbonyl-Protected Aminosiloxane Monolayers" J. Am. Chem. Soc., 2009, 131, 1513-1522. (PDF)
• K. Okrasa, C. Levy, N. Baudistel, D. Leys and J. Micklefield, "Structure and mechanism of an unusual malonate decarboxylase and related racemases", Chem. Eur. J., 2008, 14, 6609-6613.(PDF)
• B. Amir-Heidari, J. Thirlway and J. Micklefield, "Auxotrophic precursor-directed biosynthesis of nonribosomal lipopeptides with modified tryptophan residues", Org. Biomol. Chem., 2008, 6, 975-978. (PDF) Highlighted on the front cover (see right, click on image to see original)
• S. Vaidyanthan, J. S. Fletcher, N. P. Lockyer, R. Goodacre, J. Micklefield and J. C. Vickerman, "Subsurface biomolecular imaging of Streptomyces coelicolor using mass spectrometry", Anal. Chem., 2008, 80, 1942-1951.(PDF)
• R. Worthington, N. Bell and J. Micklefield, "RNA selective cross-pairing of backbone extended pyrrolidine-amide oligonucleotide mimics (bePOMs", Org. Biomol. Chem., 2008, 6, 92-103. (PDF)
• A. Powell, M. Borg B. Amir-Heidari, J. M. Neary, J. Thirlway, B. Wilkinson, C.P. Smith, J. Micklefield, “Engineered biosynthesis of nonribosomal lipopeptide antibiotics with modified fatty acid side chains” J. Am. Chem. Soc. 2007, 129, 15182-15191 (PDF)
• C. Mahlert, F.Kopp, J Thirlway, J. Micklefield, and M.A. Marahiel, "Stereospecific Enzymatic Transformation of r-Ketoglutarate to (2S,3R)-3-Methyl Glutamate during Acidic Lipopeptide Biosynthesis", J. Am. Chem. Soc., 2007, 129, 12011-12018. (PDF)
• B. Wilkinson, J. Micklefield “Mining and engineering natural-product biosynthetic pathways” Nature Chemical Biology 2007, 3, 379-386. (PDF)
• R.J. Worthington, N.M. Bell, R. Wong and J. Micklefield, "RNA-selective cross-pairing of backbone-extended pyrrolidine-amide oligonucleotide mimics (bePOMs)", Organic & Biomolecular Chemistry, 2007, 6, 92-107. (PDF)
• B. Amir-Heidari and J. Micklefield “NMR Confirmation that Tryptophan Dehydrogenation Occurs with Syn-stereochemistry During the biosynthesis of CDA in Streptomyces coelicolor” J. Org. Chem. 2007, 72, 8950-8953. (PDF)

• N. Dixon, L. S. Wong, T. Geerlings and J. Micklefield “Cellular targets of natural products” Natural Products Reports 2007, 24, 1288-1310. (PDF) Highlighted on the front cover (see right, click on image to see original)
• R. Worthington, N. Bell and J. Micklefield “RNA selective cross- pairing of backbone extended pyrrolidine-amide oligonucleotide mimics (bePOMs).” Org. Biomol. Chem. 2007 in press
  

• A. Powell, M. Al Nakeeb, B. Wilkinsonb and J. Micklefield, "Precursor-directed biosynthesis of nonribosomal lipopeptides with modified glutamate residues", Chem. Commun., 2007, 2683-2685. (PDF) Highlighted on the front cover (see left, click on image to see original)
• B. A. Heidari, J. Thirlway, J. Micklefield "Stereochemical Course of Tryptophan Dehydrogenation during Calcium-Dependent Lipopeptide Antibiotic Biosynthesis" Org. Lett. 2007, 9, 1513-1516. (PDF)
• J. Neary, A. Powell, L. Gordon C. Milne, F. Flett, B. Wilkinson, C.P. Smith and J. Micklefield "An asparagine oxygenase (AsnO) and a 3- hydroxyasparaginyl phosphotransferase (HasP) are involved in the biosynthesis of calcium dependent lipopeptide antibiotics" Microbiology. 2007, 153, 768-776. (PDF)
• S.T.H. Tan, R. Worthington, A. O’Rourke J. Morral R. G. Pritchard and J. Micklefield "Homopolymeric pyrrolidine-amide oligonucleotidemimics: Fmoc-synthesis and DNA/RNA binding properties" Org. Biomol. Chem., 2007, 5, 239-248. (PDF)

• R. Worthington, A. O’Rourke J. Morral, S.T.H. Tan, and J. Micklefield "Mixed-sequence pyrrolidine-amide oligonucleotide mimics: Boc(Z) synthesis and DNA/RNA binding properties." Org. Biomol. Chem., 2007, 5, 249-259. (PDF) Highlighted on the front cover (see right, click on image to see original)
• C. Milne, A. Powell, J. Jim, M. Al Nakeeb C. P. Smith, J. Micklefield, "Biosynthesis of the (2S,3R)-3-methyl glutamate residue of nonribosomal lipopeptides" J. Am. Chem. Soc., 2006, 128, 11250-11259. (PDF)
• A. Kahn & J. Micklefield, "Effects of stereospecific backbone methylation on the DNA/RNA binding properties of pyrrolidine-amide oligonucleotide mimics (cPOM)" Chem Commun., 2006,1436 - 1438. (PDF)
• S. Saul, S. Corr and J. Micklefield “Biotransformations in low-boiling hydrofluorocarbon solvents” Angew. Chem. Int. Ed. 2004, 43, 5519-5523. (PDF)
• J. Micklefield "Daptomycin structure and mechanism of action revealed" Chemistry & Biology, 2004, 11, 887-888. (PDF)

• L.-J. Ball, C.M. Goult, J.A. Donarski, J. Micklefield and V. Ramesh “NMR structure determination and calcium binding effects of lipopeptide antibiotic daptomycin” Org. Biomol. Chem., 2004, 2, 1872–1878. (PDF) This paper featured on the front cover of this issue of Organic and Biomolecular Chemistry (see right)
• H.B. Kim, C.P. Smith, J. Micklefield, F. Mavituna, "Metabolic flux balancing for calcium dependent antibiotic (CDA) production in Streptomyces coelicolor", Metabolic Engineering, 2004, 6, 313-325. (PDF)
• G.C. Uguru, C. Milne, M. Borg, F. Flett, C.P. Smith, J. Micklefield, "Active-Site Modifications of Adenylation Domains Lead to Hydrolysis of Upstream Nonribosomal Peptidyl Thioester Intermediates", J. Am. Chem. Soc., 2004, 126, 5032-5033. (PDF)
• H. Li, P. Williams, J Micklefield, J.M. Gardiner and G. Stephens, "A dynamic combinatorial screen for novel imine reductase activity", Tetrahedron, 2004, 60, 753–758. (PDF)
• T. H. Samuel Tan, David T. Hickman, Jordi Morral, Ian G. Beadham and Jason Micklefield, "Nucleic acid binding properties of thyminyl and adeninyl pyrrolidine-amide oligonucleotide mimics (POM)", Chem. Comm., 2004, 516-517. (PDF)
• D.T. Hickman, T.H.S. Tan, J. Morral, P.M. King, M.A. Cooper and J. Micklefield, "Design, synthesis, conformational analysis and nucleic acid hybridisation properties of thymidyl pyrrolidine-amide oligonucleotide mimics (POM)", Org. Biomol. Chem., 2003, 1, 3277-3292. (PDF)

• Z. Hojati, C. Milne, B. Harvey, L. Gordon, M. Borg, F. Flett, B.Wilkinson, P. J. Sidebottom, B. A. M. Rudd, M. A. Hayes, C. P. Smith and J. Micklefield, "Structure, Biosynthetic Origin, and Engineered Biosynthesis of Calcium-Dependent Antibiotics from Streptomyces coelicolor", Chem. Biol. 2002, 9, 1175-1187. (PDF, Preview PDF) This paper featured on the front cover of this Chemistry and Biology issue (see right)
• K. J. Fettes, N. Howard, D. T. Hickman, S. A. Adah, M. R. Player, P. F. Torrence and J. Micklefield , "Synthesis and Nucleic-Acid-Binding Properties of Sulfamide and 3´NSulfamate Modified DNA",  J. Chem. Soc., Perkin Trans. 1. 2002, 485-495. (PDF)
• D. T. Hickman, P. M. King, J. M. Slater, M. A. Cooper and J. Micklefield "Kinetically selective binding of single stranded RNA over DNA by a pyrrolidine­amide oligonucleotide mimic (POM)" Nucleosides, Nucleotides & Nucleic Acids, 2001, 20, 1169-1172.(PDF)
• J. Micklefield "Backbone Modification of Nucleic Acids: Synthesis, Structure and Therapeutic Applications." Current Medicinal Chemistry 2001, 8, 1157-1180. (PDF)
• D. T. Hickman, P. M. King, M. A. Cooper, J. M. Slater and J. Micklefield "Unusual RNA & DNA binding properties of a novel pyrrolidine­amide oligonucleotide mimic (POM)" Chem. Commun. 2000, 2251-2252. (PDF)
• K. J. Fettes, N. Howard, D. T. Hickman, S. A. Adah, M. R. Player, P. F. Torrence and J. Micklefield, "Replacement of the phosphodiester linkage in DNA with sulfamide and 3´-N-sulfamate groups " Chem. Commun., 2000, 765-766. (PDF)

Patents

• S.J.P. Saul, J. Micklefield and S. Corr "Enzymatic process for stereo-selective preparation of chemical compounds in hydrofluorocarbon solvents", PCT Int. Appl. (2004), WO2004083444. (PDF)
• J. Micklefield, A. J. Ball, S. Corr “Process for dynamic kinetic resolution (DKR) of racemic compounds in hydrofluorocarbon solvents. PCT Int. Appl. (2007), WO 2007129018.