Florian Hollfelder

Summary

Affiliation: University of Cambridge
Country: UK

Publications

  1. pmc Isothermal DNA amplification using the T4 replisome: circular nicking endonuclease-dependent amplification and primase-based whole-genome amplification
    Yolanda Schaerli
    Department of Biochemistry, University of Cambridge, Cambridge, UK
    Nucleic Acids Res 38:e201. 2010
  2. pmc In vitro affinity screening of protein and peptide binders by megavalent bead surface display
    Letizia Diamante
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, CB2 1GA Cambridge, UK
    Protein Eng Des Sel 26:713-24. 2013
  3. pmc Peptide dendrimer/lipid hybrid systems are efficient DNA transfection reagents: structure--activity relationships highlight the role of charge distribution across dendrimer generations
    Albert Kwok
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, United Kingdom
    ACS Nano 7:4668-82. 2013
  4. doi request reprint Picoliter cell lysate assays in microfluidic droplet compartments for directed enzyme evolution
    Balint Kintses
    Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, UK
    Chem Biol 19:1001-9. 2012
  5. doi request reprint Droplets as reaction compartments for protein nanotechnology
    Sean R A Devenish
    Department of Biochemistry, University of Cambridge, Cambridge, UK
    Methods Mol Biol 996:269-86. 2013
  6. doi request reprint Efficient catalytic promiscuity in an enzyme superfamily: an arylsulfatase shows a rate acceleration of 10(13) for phosphate monoester hydrolysis
    Luis F Olguin
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, United Kingdom
    J Am Chem Soc 130:16547-55. 2008
  7. doi request reprint Microdroplets in microfluidics: an evolving platform for discoveries in chemistry and biology
    Ashleigh B Theberge
    Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge, CB2 1EW UK
    Angew Chem Int Ed Engl 49:5846-68. 2010
  8. doi request reprint Simultaneous determination of gene expression and enzymatic activity in individual bacterial cells in microdroplet compartments
    Jung uk Shim
    Department of Biochemistry, University of Cambridge, Cambridge, CB2 1GA, United Kingdom
    J Am Chem Soc 131:15251-6. 2009
  9. pmc A single mutation in the core domain of the lac repressor reduces leakiness
    Pietro Gatti-Lafranconi
    Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, UK
    Microb Cell Fact 12:67. 2013
  10. pmc An efficient, multiply promiscuous hydrolase in the alkaline phosphatase superfamily
    Bert van Loo
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, United Kingdom
    Proc Natl Acad Sci U S A 107:2740-5. 2010

Collaborators

Detail Information

Publications50

  1. pmc Isothermal DNA amplification using the T4 replisome: circular nicking endonuclease-dependent amplification and primase-based whole-genome amplification
    Yolanda Schaerli
    Department of Biochemistry, University of Cambridge, Cambridge, UK
    Nucleic Acids Res 38:e201. 2010
    ....
  2. pmc In vitro affinity screening of protein and peptide binders by megavalent bead surface display
    Letizia Diamante
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, CB2 1GA Cambridge, UK
    Protein Eng Des Sel 26:713-24. 2013
    ....
  3. pmc Peptide dendrimer/lipid hybrid systems are efficient DNA transfection reagents: structure--activity relationships highlight the role of charge distribution across dendrimer generations
    Albert Kwok
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, United Kingdom
    ACS Nano 7:4668-82. 2013
    ..In the future, this platform may be tuned further and coupled to cell-targeting moieties to enhance transfection and cell specificity...
  4. doi request reprint Picoliter cell lysate assays in microfluidic droplet compartments for directed enzyme evolution
    Balint Kintses
    Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, UK
    Chem Biol 19:1001-9. 2012
    ..The detection of subtle differences in a larger number of screened clones provides the combination of high sensitivity and high-throughput needed to rescue a stalled directed evolution experiment and make it viable...
  5. doi request reprint Droplets as reaction compartments for protein nanotechnology
    Sean R A Devenish
    Department of Biochemistry, University of Cambridge, Cambridge, UK
    Methods Mol Biol 996:269-86. 2013
    ....
  6. doi request reprint Efficient catalytic promiscuity in an enzyme superfamily: an arylsulfatase shows a rate acceleration of 10(13) for phosphate monoester hydrolysis
    Luis F Olguin
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, United Kingdom
    J Am Chem Soc 130:16547-55. 2008
    ....
  7. doi request reprint Microdroplets in microfluidics: an evolving platform for discoveries in chemistry and biology
    Ashleigh B Theberge
    Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge, CB2 1EW UK
    Angew Chem Int Ed Engl 49:5846-68. 2010
    ..Examples are presented to show how compartmentalization, monodispersity, single-molecule sensitivity, and high throughput have been exploited in experiments that would have been extremely difficult outside the microfluidics platform...
  8. doi request reprint Simultaneous determination of gene expression and enzymatic activity in individual bacterial cells in microdroplet compartments
    Jung uk Shim
    Department of Biochemistry, University of Cambridge, Cambridge, CB2 1GA, United Kingdom
    J Am Chem Soc 131:15251-6. 2009
    ..By collecting data from many individual cells, the distribution of activities in a cell is quantified and the difference in activity between two AP mutants is measured...
  9. pmc A single mutation in the core domain of the lac repressor reduces leakiness
    Pietro Gatti-Lafranconi
    Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, UK
    Microb Cell Fact 12:67. 2013
    ..However, the leakiness of LacI that is essential for the natural function of the lac operon leads to an increased energetic burden, and potentially toxicity, in heterologous protein production...
  10. pmc An efficient, multiply promiscuous hydrolase in the alkaline phosphatase superfamily
    Bert van Loo
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, United Kingdom
    Proc Natl Acad Sci U S A 107:2740-5. 2010
    ..PMH is thus one of the most promiscuous hydrolases described to date. The functional links between superfamily activities can be presumed to have played a role in functional evolution by gene duplication...
  11. ncbi request reprint An experimental framework for improved selection of binding proteins using SNAP display
    Gillian Houlihan
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, UK MedImmune Ltd, Milstein Building, Granta Park, Cambridge, CB1 6GH, UK
    J Immunol Methods 405:47-56. 2014
    ..These data establish SNAP display as an in vitro protein engineering tool for isolating protein binders and provide a framework for troubleshooting affinity selections. ..
  12. ncbi request reprint A simple method to evaluate the biochemical compatibility of oil/surfactant mixtures for experiments in microdroplets
    Miriam Kaltenbach
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, CB2 1GA, Cambridge, UK
    Lab Chip 12:4185-92. 2012
    ..This methodology will serve as a quantitative test for the rapid optimization of droplet-based experiments such as in vitro protein expression or enzymatic assays...
  13. doi request reprint Controlling the contents of microdroplets by exploiting the permeability of PDMS
    Jung uk Shim
    Department of Chemistry, University of Cambridge, Cambridge, UK
    Lab Chip 11:1132-7. 2011
    ..Likewise quorum sensing in bacteria was triggered by delivery of the auto-inducer N-(3-oxododecanoyl)-l-homoserine lactone (OdDHL) through the PDMS membrane of the device...
  14. doi request reprint A double droplet trap system for studying mass transport across a droplet-droplet interface
    Yunpeng Bai
    Melville Laboratory for Polymer Synthesis, Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge, UKCB2 1EW
    Lab Chip 10:1281-5. 2010
    ....
  15. doi request reprint Monitoring a reaction at submillisecond resolution in picoliter volumes
    Ansgar M Huebner
    Department of Biochemistry, University of Cambridge, Cambridge, United Kingdom
    Anal Chem 83:1462-8. 2011
    ..Our analysis makes kinetics accessible that are several times faster than techniques that have to allow time for mixing...
  16. doi request reprint Controlling the retention of small molecules in emulsion microdroplets for use in cell-based assays
    Fabienne Courtois
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge, CB2 1GA, UK
    Anal Chem 81:3008-16. 2009
    ....
  17. doi request reprint Continuous-flow polymerase chain reaction of single-copy DNA in microfluidic microdroplets
    Yolanda Schaerli
    Department of Biochemistry and Department of Chemistry, University of Cambridge, Cambridge, UK
    Anal Chem 81:302-6. 2009
    ..This device holds promise for convenient integration with other microfluidic devices and adds a critical missing component to the laboratory-on-a-chip toolkit...
  18. pmc Fluid phase endocytosis contributes to transfection of DNA by PEI-25
    Hansjörg Hufnagel
    Department of Biochemistry, University of Cambridge, Cambridge, UK
    Mol Ther 17:1411-7. 2009
    ....
  19. doi request reprint An integrated cell culture lab on a chip: modular microdevices for cultivation of mammalian cells and delivery into microfluidic microdroplets
    Hansjörg Hufnagel
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge, UK CB2 1GA
    Lab Chip 9:1576-82. 2009
    ..The procedures developed for this modular device layout further demonstrate that commonly used methods in cell biology involving mammalian cells can be reliably scaled down to allow single cell investigations in picolitre volumes...
  20. pmc A fully unsupervised compartment-on-demand platform for precise nanoliter assays of time-dependent steady-state enzyme kinetics and inhibition
    Fabrice Gielen
    Department of Biochemistry, University of Cambridge, Cambridge, United Kingdom
    Anal Chem 85:4761-9. 2013
    ..In both cases, the kinetic parameters obtained in droplets were identical within error to values obtained in titer plates, despite a >10(4)-fold volume reduction, from micro- to nanoliters. ..
  21. doi request reprint Development of quantitative cell-based enzyme assays in microdroplets
    Ansgar Huebner
    Department of Biochemistry, University of Cambridge, UK
    Anal Chem 80:3890-6. 2008
    ..Studies on wild type and a mutant enzyme successfully demonstrated the feasibility of using microfluidic droplets to provide time-resolved kinetic measurements...
  22. doi request reprint Directed evolution of a histone acetyltransferase--enhancing thermostability, whilst maintaining catalytic activity and substrate specificity
    Hans Leemhuis
    Department of Biochemistry, University of Cambridge, UK
    FEBS J 275:5635-47. 2008
    ..The directed evolution approach described in the present study is easily adapted to other histone modifying enzymes, requiring only appropriate peptide substrates and antibodies, which are available from commercial suppliers...
  23. doi request reprint The potential of microfluidic water-in-oil droplets in experimental biology
    Yolanda Schaerli
    Department of Biochemistry, University of Cambridge, Cambridge, UK
    Mol Biosyst 5:1392-404. 2009
    ..This review describes the current state-of-the-art, notes salient features of successful experiments and extrapolates in the direction of more highly integrated systems...
  24. doi request reprint Simultaneous measurement of reactions in microdroplets filled by concentration gradients
    Nicolae Damean
    Department of Chemistry, University of Cambridge, Cambridge, CB2 1EW, UK
    Lab Chip 9:1707-13. 2009
    ..e. channels having identical hydraulic resistance) is exploited and developed. As a demonstration of the PmicroD technology, an enzyme activity assay was carried out and the steady-state kinetic constants were determined...
  25. doi request reprint Combining medium effects and cofactor catalysis: metal-coordinated synzymes accelerate phosphate transfer by 10(8)
    Frédéric Avenier
    Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, UK
    Chemistry 15:12371-80. 2009
    ..These characteristics render synzyme Zn(II)-CD6 an efficient enzyme model that reflects enzyme-like properties in a wide range of features...
  26. doi request reprint SNAP display: in vitro protein evolution in microdroplets
    Miriam Kaltenbach
    Department of Biochemistry, University of Cambridge, Cambridge, UK
    Methods Mol Biol 805:101-11. 2012
    ..This method will prove a useful addition to the array of in vitro display systems, distinguished by the stability of DNA as the coding nucleic acid and the covalent link between gene and protein...
  27. doi request reprint Kinetic analysis of beta-phosphoglucomutase and its inhibition by magnesium fluoride
    Marko Golicnik
    Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, United Kingdom
    J Am Chem Soc 131:1575-88. 2009
    ..The preference for TSA formation when fluoride is present, and the hydrolysis of substrates when it is not, rules out the formation of a stable pentavalent phosphorane intermediate in the active site of betaPGM...
  28. doi request reprint Static microdroplet arrays: a microfluidic device for droplet trapping, incubation and release for enzymatic and cell-based assays
    Ansgar Huebner
    Department of Biochemistry, University of Cambridge, 80, Tennis Court Road, Cambridge, UK CB2 1GA
    Lab Chip 9:692-8. 2009
    ..coli cells and enzymatic reactions. We also demonstrate that trapped droplets may be recovered from the microfluidic array for further processing...
  29. pmc An efficient method to assemble linear DNA templates for in vitro screening and selection systems
    Viktor Stein
    Department of Biochemistry, University of Cambridge, CB2 1GA, Cambridge, UK
    Nucleic Acids Res 37:e122. 2009
    ..This fast and robust method should find widespread application in directed evolution for the assembly of gene libraries and the regeneration of linear DNA templates between successive screening and selection cycles...
  30. doi request reprint Flexibility and reactivity in promiscuous enzymes
    Pietro Gatti-Lafranconi
    Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, UK
    Chembiochem 14:285-92. 2013
    ..This review analyses the roles structural flexibility and chemical reactivity play in the catalytic mechanism of selected enzymes...
  31. doi request reprint Ultrarapid generation of femtoliter microfluidic droplets for single-molecule-counting immunoassays
    Jung uk Shim
    Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge, UK, CB2 1EW
    ACS Nano 7:5955-64. 2013
    ..This work demonstrates a highly flexible and sensitive diagnostic platform that exploits extremely high-speed generation of monodisperse femtoliter droplets for the counting of individual analyte molecules. ..
  32. ncbi request reprint A covalent chemical genotype-phenotype linkage for in vitro protein evolution
    Viktor Stein
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, CB2 1GA, Cambridge, UK
    Chembiochem 8:2191-4. 2007
  33. doi request reprint Directed evolution of a gatekeeper domain in nonribosomal peptide synthesis
    Benoit Villiers
    Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, UK
    Chem Biol 18:1290-9. 2011
    ..These results establish directed evolution as a method to change gatekeeper domain specificity and suggest that adaptation of modules in combinatorial biosynthesis is achievable with few mutations during evolution...
  34. doi request reprint Assembling linear DNA templates for in vitro transcription and translation
    Viktor Stein
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, UK
    Methods Mol Biol 815:67-78. 2012
    ..This methodology will be useful for repeated cycles of expression and in vitro selection, in which gene libraries are repeatedly assembled and their products and templates regenerated...
  35. doi request reprint SNAP dendrimers: multivalent protein display on dendrimer-like DNA for directed evolution
    Miriam Kaltenbach
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, CB2 1GA Cambridge, UK
    Chembiochem 12:2208-16. 2011
    ..These data suggest that multivalent SNAP dendrimers have the potential to facilitate the selection of protein binders especially during early rounds of directed evolution, allowing a larger diversity of candidate binders to be recovered...
  36. ncbi request reprint New genotype-phenotype linkages for directed evolution of functional proteins
    Hans Leemhuis
    Department of Biochemistry, Cambridge University, Cambridge CB2 1GA, UK
    Curr Opin Struct Biol 15:472-8. 2005
    ..New approaches such as mRNA display, ribosome display and in vitro compartmentalisation have complementary areas of application in selections for binding or catalysis...
  37. ncbi request reprint Kinetic and computational evidence for an intermediate in the hydrolysis of sulfonate esters
    Ann C Babtie
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, UK
    Org Biomol Chem 10:8095-101. 2012
    ..The emerging scenario can be explained by the combined effect of a strong nucleophile with a poor leaving group that compel a usually concerted reaction to favour a stepwise process...
  38. doi request reprint Microdroplets: a sea of applications?
    Ansgar Huebner
    Department of Chemistry, Lensfield Road, Cambridge, UKCB2 1EW
    Lab Chip 8:1244-54. 2008
    ..Furthermore, we discuss key fields of use in which such systems may make a significant impact, with particular emphasis on novel applications in the biological and physical sciences...
  39. doi request reprint Microfluidic droplets: new integrated workflows for biological experiments
    Balint Kintses
    Department of Biochemistry, University of Cambridge, United Kingdom
    Curr Opin Chem Biol 14:548-55. 2010
    ..Directed evolution, diagnostics and compound screening are areas in which the first steps are being taken toward the long-term goal of transforming the way we design and carry out experiments...
  40. doi request reprint Relating chemical and biological diversity space: a tunable system for efficient gene transfection
    Liisa D Van Vliet
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge, CB2 1GA, UK
    Chembiochem 9:1960-7. 2008
    ....
  41. doi request reprint A new member of the alkaline phosphatase superfamily with a formylglycine nucleophile: structural and kinetic characterisation of a phosphonate monoester hydrolase/phosphodiesterase from Rhizobium leguminosarum
    Stefanie Jonas
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB21GA, UK
    J Mol Biol 384:120-36. 2008
    ..coli alkaline phosphatase and Pseudomonas aeruginosa arylsulfatase. R/PMH represents a further example of conservation of the overall structure and mechanism within the alkaline phosphatase superfamily...
  42. pmc One in a million: flow cytometric sorting of single cell-lysate assays in monodisperse picolitre double emulsion droplets for directed evolution
    Anastasia Zinchenko
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, U K
    Anal Chem 86:2526-33. 2014
    ....
  43. ncbi request reprint SNAP display - an in vitro method for the selection of protein binders
    Gillian Houlihan
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, UK
    Curr Pharm Des 19:5421-8. 2013
    ..This review describes the principles of SNAP display and extrapolates its scope for in vitro selections. ..
  44. doi request reprint What makes an enzyme promiscuous?
    Ann Babtie
    Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, UK
    Curr Opin Chem Biol 14:200-7. 2010
    ....
  45. ncbi request reprint Mapping the limits of substrate specificity of the adenylation domain of TycA
    Benoit R M Villiers
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge, UK
    Chembiochem 10:671-82. 2009
    ..The specificity patterns observed and the quantification of alternative activities provide a basis for exploring possible paths for the future directed evolution of A-domain specificity...
  46. doi request reprint Efficient catalytic promiscuity for chemically distinct reactions
    Ann C Babtie
    Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, UK
    Angew Chem Int Ed Engl 48:3692-4. 2009
    ....
  47. doi request reprint The human histone acetyltransferase P/CAF is a promiscuous histone propionyltransferase
    Hans Leemhuis
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, UK
    Chembiochem 9:499-503. 2008
  48. doi request reprint An integrated device for monitoring time-dependent in vitro expression from single genes in picolitre droplets
    Fabienne Courtois
    Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge CB2 1EW, UK
    Chembiochem 9:439-46. 2008
    ..The high efficiency allows protein expression from a single molecule of DNA template, creating "monoclonal droplets" in which genotype and phenotype are combined in one emulsion compartment...
  49. ncbi request reprint Molecular recognition of DNA by rigid [N]-polynorbornane-derived bifunctional intercalators: synthesis and evaluation of their binding properties
    Liisa D Van Vliet
    Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, United Kingdom
    J Med Chem 50:2326-40. 2007
    ..The flexibility of the linker connecting the rings to the scaffold, although less influential, could affect the strength and character of the DNA binding...
  50. doi request reprint Efficient, crosswise catalytic promiscuity among enzymes that catalyze phosphoryl transfer
    Mark F Mohamed
    Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, EU, UK
    Biochim Biophys Acta 1834:417-24. 2013
    ..This article is part of a Special Issue entitled: Chemistry and mechanism of phosphatases, diesterases and triesterases...