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Department of Biochemistry

Florian Hollfelder

Mechanism in Chemistry and Biology

Nature has evolved the most amazing functional biomolecules and we want to know how they work. Enzymes are the all-purpose catalysts that make the Chemistry of Life run smoothly and efficiently, under the mildest, ‘greenest’ conditions – and protein binders are involved in governing many biological processes.

This group is interested in gaining a fundamental understanding of the principles responsible for molecular recognition processes in chemistry and biology. We probe whether these principles enable us to describe, manipulate and ultimately make functional molecules. Using an eclectic mix of techniques (including kinetic measurements, organic synthesis, biophysical analysis and microengineering) we extend the mechanistic lessons learned to potential applications in biotechnology, chemistry and medicine. In particular we are excited about the potential of pico- to nanolitre mircrodroplets to provide a quantitative high-throughput system that provides insight not only into one reaction at a time, but - by way of massive multiplexing - lets us study millions of samples in one go.

Lab members: Chloé Bleuez, Lise BoursinhacEleanor Campbell, Joachim De Jonghe, Anna Lena Ellermann, Katrin Fischer, Thomas Fryer, Michael Herger, Josephin Holstein, Tomasz Kaminski, Miriam Klaus, Philipp Knyphausen, Wolfgang Koch, Timo Kohler, Simon Ladevèze, Philip Mair, Alisa MaringKatharina Neufeld, Stefanie Neun, Maya Petek, Mariana Rangel Pereira, Joel Rogers, Remkes Scheele, David Schnettler, Liisa Van Vliet, Paul Zurek.

Visit Hollfelder group website

Key publications:

• Miton, C. M.; Jonas, S.; Fischer, G; Duarte, F.; Mohamed, M.F.; van Loo, B.;  Kintses, B.; Kamerlin,, S. C. L.; Tokuriki , N.; Hyvönen, M.; Hollfelder, F.  Evolutionary repurposing of a sulfatase: a new Michaelis complex leads to efficient transition state charge offset Proc. Natl. Acad. Sci USA 2018, 115(31):E7293-E7302. doi: 10.1073/pnas.1607817115.

• van Loo, B.; Schober, M.; Valkov, E.; Heberlein, M.; Bornberg-Bauer, E.; Faber, K.; Hyvönen; Hollfelder, F.  Structural and Mechanistic Analysis of the Choline Sulfatase from Sinorhizobium melliloti: A Class I Sulfatase Specific for an Alkyl Sulfate Ester J Mol Biol 2018, 430(7):1004-1023. doi:10.1016/j.jmb.2018.02.010.

• Bayer, C.; van Loo, B.; Hollfelder, F. Specificity effects of amino acid substitutions in promiscuous hydrolases – context-dependence of catalytic residue contributions to local fitness landscapes in nearby sequence space; ChemBioChem 2017,  doi:10.1002/cbic.201600657

• Mair, P.; Gielen, F.; Hollfelder, F., Exploring sequence space in search of functional enzymes using microfluidic droplets. Curr Opin Chem Biol 2017, 137-144, doi: 10.1016/j.cbpa.2017.02.018

• Gielen F, Hours R, et al. Ultrahigh-throughput-directed enzyme evolution by absorbance-activated droplet sorting (AADS). Proc. Natl Acad. Sci USA 2016, 113(47), E7383-E7389. Read online

• Campbell, E.; Kaltenbach, M.; Correy, G. J.; Carr, P. D.; Porebski, B. T.; Livingstone, E. K.; Afriat-Jurnou, L.; Buckle, A. M.; Weik, M.; Hollfelder, F.; Tokuriki, N.; Jackson, C. J., The role of protein dynamics in the evolution of new enzyme function. Nature Chem Biol 2016, 12 (11), 944-950.

• Colin PY, et al. Ultrahigh-throughput discovery of promiscuous enzymes by picodroplet functional metagenomics. Nature Communications 2015, 6, 10008. Read online

• Colin PY, Zinchenko A, Hollfelder F. Enzyme engineering in biomimetic compartments. Curr Opin Struct Biol 2015, 33, 42-51. Read online

• Shafee T, Gatti-Lafranconi P, Minter R, Hollfelder F. Handicap-Recover Evolution Leads to a Chemically Versatile, Nucleophile-Permissive Protease. Chembiochem 2015, 16(13), 1866–1869. Read online

• Fischlechner M, Schaerli Y, Mohamed MF, Patil S, Abell C & Hollfelder F. Microfluidic Gel-Shell Beads: Evolution of Catalysts Caged in Biomimetic Compartments. Nature Chemistry 2014, 6(9):791-6. Read online

• Diamante, L.; Gatti-Lafranconi, P.; Schaerli, Y.; Hollfelder, F., In vitro affinity screening of protein and peptide binders by megavalent bead surface display. Protein Eng Des Sel 2013, 26(10):713-24. Read online

• Mohamed, M.F. & Hollfelder, F. Efficient, crosswise catalytic promiscuity among enzymes that catalyze phosphoryl transfer. Biochim Biophys Acta 2013, 1834, 417-2. Read online

• Kwok, A., Eggimann, G.A., Reymond, J.L., Darbre, T. & Hollfelder, F. Peptide dendrimer/lipid hybrid systems are efficient DNA transfection reagents: structure-activity relationships highlight the role of charge distribution across dendrimer generations. ACS Nano 2013, 7, 4668-82. Read online

• Gielen, F.; van Vliet, L.; Koprowski, B. T.; Devenish, S. R.; Fischlechner, M.; Edel, J. B.; Niu, X.; Demello, A. J.; Hollfelder, F., A Fully Unsupervised Compartment-on-Demand Platform for Precise Nanoliter Assays of Time-Dependent Steady-State Enzyme Kinetics and Inhibition. Anal Chem 2013, 85(9):4761-9. Read online

• Kintses, B.; Hein, C.; Mohamed, M. F.; Fischlechner, M.; Courtois, F.; Lainé, C.; Hollfelder, F. Chemistry & Biology 2012, Picoliter cell lysate assays in microfluidic droplet compartments for directed enzyme evolution, 19(8):1001-9. Read online

• Villiers, B. R.; Hollfelder, F., Directed evolution of a gatekeeper domain in nonribosomal peptide synthesis. Chemistry & Biology 2011,18(10):1290-9. Read online

• van Loo, B.; Jonas, S.; Babtie, A. C.; Benjdia, A.; Berteau, O.; Hyvönen, M.; Hollfelder, F., An efficient, multiply promiscuous hydrolase in the alkaline phosphatase superfamily. Proc. Natl Acad. Sci USA 2010, 107 (7), 2740-5. Read online

• Golicnik, M.; Olguin, L. F.; Feng, G.; Baxter, N. J.; Waltho, J. P.; Williams, N. H.; Hollfelder, F., Kinetic analysis of beta-phosphoglucomutase and its inhibition by magnesium fluoride. J Am Chem Soc 2009, 131 (4), 1575-88. Read online

• Shim, J. U.; Olguin, L. F.; Whyte, G.; Scott, D.; Babtie, A.; Abell, C.; Huck, W. T.; Hollfelder, F., Simultaneous determination of gene expression and enzymatic activity in individual bacterial cells in microdroplet compartments. J Am Chem Soc 2009, 131 (42), 15251-6. Read online