BIOINFORMATICS AND MOLECULAR MODELING IN PROTEIN STRUCTURE, FUNCTION, AND ENGINEERING
Project PI: Dr. Xevi Biarnés
Current research addresses the development and application of computational biology methods to the fundamental understanding of the molecular mechanisms governing key processes in the cell life cycle and the ways to interact with them to achieve applications in biomedicine and biotechnology. In particular:
– the development of computer assisted protocols to assist experimental protein engineering for the design of new biocatalysts by means of structural bioinformatics, computational genomics and atomistic simulations.
– the study of conformational changes of biological macromolecules such as protein folding, molecular recognition events such as protein-protein or protein-ligand interactions, and biological reactivity such as enzymatic catalysis.
We rely on the use of high-performance computing infrastructures, such as MareNostum (Spain), Cineca (Italy), Abaco (IQS) for the execution of our research projects.
a) BindScan: a computational algorithm to predict hot-spots for experimental directed evolution of proteins
In the perspective of using enzymes as biocatalysts for the production of non-natural compounds, the starting substrates are usually non-natural as well. Since the enzyme-substrate interactions are not naturally optimized in such cases, it is expected to be much room for the improvement of the catalytic efficiency. Modifications on enzyme structure may lead to the adaptation of enzyme cavities to the binding of new substrates, and to the establishment of new specific enzyme-substrate interactions that may improve its catalytic efficiency. Identifying the regions of the enzyme structure most sensible to the binding of a given non-natural substrate is crucial for the successful redesign of the enzyme.
We have developed an algorithm (named “BindScan”) that exhaustively casts all the positions on a given protein sequence by individually mutating each position and measuring the effect on the binding and reactivity to a given compound. The only requirements for such algorithm are the three-dimensional structures of both the test compound and the enzyme to be optimized. The algorithm is able to identify hot-spots along the protein sequence for which an improve of catalytic efficiency towards the tested compound is expected upon randomization. We have already successfully applied this algorithm to different test cases [1], and we are currently working at improving the theoretical background and computer efficiency of the algorithm.
Rational design of a highly efficient transglycosylating enzyme predicted by BindScan protocol (Bissaro et al. ACSCatal 2015).
Selected publications:
Molecular Design of Non-Leloir Furanose-Transferring Enzymes from an α-L-Arabinofuranosidase: A Rationale for the Engineering of Evolved Transglycosylases.
B. Bissaro, J. Durand, X.Biarnés., A. Planas, P. Monsan, R. Fauré, M. J. O’Donohue.
ACS Catalysis 5, 4598-4611 (2015). Abstract. http://dx.doi.org/10.1021/acscatal.5b00949
A single point mutation alters the hydrolysis/transglycosylation partition, significantly enhancing the synthetic capability of an endo-glycoceramidase.
J. Durand, X..Biarnés, L. Watterlot, C. Bonzom, V. Borsenberger, A. Planas, S. Bozonnet, M.J. O’Donohue, R. Fauré.
ACS Catalysis 6, 8264–8275 (2016). Abstract. http://dx.doi.org/10.1021/acscatal.6b02159
Enzymatic production of defined chitosan oligomers with a specific pattern of acetylation using a combination of chitin oligosaccharide deacetylases.
S.N. Hamer, S. Cord-Landwehr, X. Biarnés, A. Planas, H. Waegeman, B.M. Moerschbacher, S. Kolkenbrock.
Scientific Reports 5, article 8716 (2015). Abstract. http://dx.doi.org/10.1038/srep08716
b) Simulation of protein conformational changes in Carbohydrate Active Enzymes
Proteins are not static objects. Besides the natural trafficking of protein biosynthesis and degradation, proteins adopt multiple different conformations which can range from small changes in side-chains orientations, to big rearrangements of secondary structure elements. These changes in geometry often have important implication in protein function. The relationships between protein dynamics and protein function are not yet widely understood for every family of enzymes. Neither the protein folding problem has been solved so far. From one hand, traditional structure determination methodologies (such as X-Ray, or even NMR) capture only a tiny portion of such conformations. On the other, short-lived structural conformers can appear in solution and may be relevant to protein function. Yet structural information for some of these slow conformational transitions are currently being disclosed thanks to the emerging Cryo-EM technology.
We are currently addressing the study of protein folding and protein conformational changes by means of computer simulation techniques based on molecular dynamics. In particular, we apply enhanced sampling techniques, such as Metadynamics. Metadynamics is a recent computer simulation approach that allows simulating slow processes such as protein folding and protein-ligand binding, for which we have accumulated long expertise. We select case studies among enzymes currently under characterization in the laboratory and collaborations with other laboratories. These include, but are not limited to, carbohydrate active enzymes, such as glycosyltransferases which are known to be highly flexible and to adopt several substrate-induced conformations.
Motion of subsite-capping loops in Vibrio cholerae chitin deacetylase predicted by Molecular Dynamics simulations.
Selected publications:
Essential mycoplasma glycolipid synthase adheres to the cell membrane by means of an amphipathic helix.
J. Romero-García, X. Biarnés, A. Planas.
Scientific Reports 9, 7085 (2019). Abstract. https://doi.org/10.1038/s41598-019-42970-9
Oxazoline or oxazolinium ion? The protonation state and conformation of the reaction intermediate of chitinase enzymes revisited.
J. Coines, M. Alfonso-Prieto, X. Biarnés, A. Planas, C. Rovira.
Chemistry A European Journal 24, 19258-19265 (2018). Abstract. http://dx.doi.org/10.1002/chem.201803905
Structural Snapshots and Loop Dynamics Along the Catalytic Cycle of Glycosyltransferase GpgS.
D. Albesa-Jové, J. Romero-García, E. Sancho-Vaello, F.X. Contreras, A. Rodrigo-Unzueta, N. Comino, A. Carreras-González, P. Arrasate, S. Urresti, X. Biarnés, A. Planas, M.E. Guerin.
Structure 25, 1034-1044 (2017). Abstract. http://dx.doi.org/10.1016/j.str.2017.05.009
A natural ternary complex trapped in crystal reveals the catalytic mechanism of a retaining glycosyltransferase.
D. Albesa-Jové, M.F. Mendoza, A. Rodrigo-Unzueta, F. Gomollón Bel, J. Cifuente, S. Urresti, N. Comino, H. Gómez, J..Romero-García, J.M. Lluch, E. Sancho-Vaello, X. Biarnés, A. Planas, P. Merino, L. Masgrau, M.E. Guerin.
Angewandte Chemie Int. Ed. 54, 9898-9902 (2015). Abstract. http://dx.doi.org/10.1002/anie.201504617
The Conformational Free Energy Landscape of β-D-Glucopyranose. Implications for Substrate Preactivation in β-Glucoside Hydrolases.
X. Biarnés, A. Ardevol, A. Planas, C. Rovira, A. Laio, M. Parrinello.
Journal of the American Chemical Society 129, 10686-10693 (2007). https://doi.org/10.1021/ja068411o
c) Evolution of protein sequence, structure and function in Carbohydrate Active Enzymes.
The most recent advances in genome sequencing are delivering huge amounts of genomic data of many different organisms. This provides the basis for understanding living systems as a whole from a molecular perspective. However, genome sequences alone are not informative. In the exciting post-genomic era, many different flavors of experimental information are routinely being incorporated into raw genomic data from the fields of genomics, transcriptomics, proteomics and metabolomics: gene regulation, differential gene expression, natural variations, clinical variants, gene function, protein domains organization, protein structure, protein function, metabolic pathways, … Bioinformatics plays a key role here in the integration of this milieu of data into comprehensible information that can be used by researchers from different fields to elaborate and corroborate their hypothesis.
This research line addresses the integration of different comparative genomics tools, data mining, protein function classification and protein structure to address the characterization of families of proteins of relevant interest for biotechnological applications. We are currently developing new automatic sequence, function, and structure analysis pipelines. These are being integrated into a unified comprehensive framework to assist in the understanding of evolutionary relationships between protein sequence, structure and function in detail. In parallel, this framework is also feeding the development of our computational assisted methods for protein design (see research line a). We select case studies to apply this framework among the families of enzymes under characterization at the Laboratory of Biochemistry and collaborations with other laboratories. These include, but are not limited to, carbohydrate active enzymes, such as chitin deacetylases and glycosyltransferases which have evolved in different species to be active on different types of substrates. (see Figure).
Subsite-Capping model proposed for the mode of action of chitin deacetylsaes family of enzymes. Model elaborated by means of hidden Markov modelling and structural alignment. (Grifoll-Romero et al. J.Biol.Chem 2019, Andrés et al. Angew.Chem.Int.Ed 2014)
Selected publications:
Structure-function relationships underlying the dual N-acetylmuramic and N-acetylglucosamine specificities of the peptidoglycan deacetylase PdaC from Bacillus subtilis.
L.Grifoll-Romero, M.A. Sainz-Polo, D. Albesa-Jové, M.E. Guerin, X. Biarnés, A. Planas.
Journal of Biological Chemistry 294, 19066-80 (2019). Abstract. https://doi.org/10.1074/jbc.RA119.009510
Structural-functional analysis reveals a specific domain organization in family GH20 hexosaminidases.
C. Val-Cid, X. Biarnés, M. Faijes, A. Planas.
PLOS One 10(5): e0128075 (2015). Abstract. http://dx.doi.org/10.1371/journal.pone.0128075
Structural basis of chitin oligosaccharide deacetylation.
E. Andrés, D. Albesa‐Jové, X. Biarnés, B.M. Moerschbacher, M.E. Guerin, A. Planas.
Angewandte Chemie International Edition 53, 6882-6887 (2014). https://doi.org/10.1002/anie.201400220
Structure-function features of a Mycoplasma glycolipid synthase derived from structural data integration, molecular simulations, and mutational analysis.
J. Romero-García, C. Francisco, X. Biarnés, A. Planas.
PLoS One 8, e81990 (2013). https://doi.org/10.1371/journal.pone.0081990