Google-Scholar Profile In one sentence, I am perhaps best described as a biochemist turned structural bioinformatician turned experimental synthetic biologist. All along, my long-term interests have been the interplay of protein dynamics, function and interactions. At present, I try to recombine modular domains into synthetic proteins and artificial protein "circuits". Such protein circuits hold promise both for exciting new diagnostic as well as therapeutic applications. Building new protein systems from scratch is also a reality check, that is, the best test of whether or not we understand how (natural) protein signalling actually works.
For my thesis, I examined different processes that put proteins on the edge of moving from one global state to another. For most of their time, proteins wiggle their atoms around some equilibrium average structure. But, when "the action is on", at the moment of mechanical stress, binding, or [insert your process of interest] these benign structure fluctuations can, as I showed, turn into decisive forces. In a first example, our study of spectrin repeats described how structural flexibility in single repeats might eventually contribute to the elasticity of red blood cells. My research then focused on the interplay between structural dynamics and protein-protein recognition and, again, structural fluctuations turned out to be a major factor determining the specificity and stability of protein-protein complexes. (Experimental studies later corroborated our computational results).
I also dedicate some time to infrastructure projects:
- Biskit, a python platform for structural bioinformatics
- BrickIt, Open source biobrick management, now reloaded as Rotten Microbes
- Evoware/py, a Python interface for Tecan robotic liquid handling
- SBOL, Standardization in Synthetic Biology. I am currently serving as an SBOL Editor.
For many years now, I have been involved with the iGEM competition and other synbio teaching activities. For example:
- as advisor to the Freiburg-Software team (2009) who developed a platform for real-time collaboration among synthetic biologists
- as advisor to the Seattle robotics "team" (2009) mostly driven by 11-year-old Gabriel who built a liquid-handling robot from original Lego pieces.
- giving a one week intensive introduction to synthetic biology at Buenos Aires University (2011 and 2013)
- as co-organizer of the first EMBO Global Exchange Lecture Course in Latin America, which drew an international crowd of students and leading synthetic biologists from Europe and the US. This course also helped kick starting Argentina's first iGEM team (2012).
- Since 2014, I have been chairing or co-chairing the committee overseeing the iGEM software track.
And here is a summary of my more official past and present scientific projects:
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senior research associate (2013-2015) |
|---|---|
| Automated DNA Assembly | |
| IRIC, University of Montreal | |
| Talk | A Field Guide to Automated Cloning (COMBINE, October 2015, Salt Lake City, Utah) [Slides] |
| Poster | Cellular and Molecular Biotechnology, 12/2015, Paris [view] |
| Abstract |
The assembly of larger DNA constructs, often from a mix of
gene synthesized and in-house fragments, is the starting
point for most synthetic biology projects. Despite major
technical advances, DNA assembly remains a bottleneck in
many laboratories.
We have implemented a robotic synthetic biology workstation that automates the complete multi-fragment DNA assembly work flow. This includes fragment PCR setup, cleanup, Gibson assembly, transformation, spreading on standard microplates, robust colony picking, colony PCR, DNA miniprep and auxiliary steps. Usability and user-adoption is facilitated by a strictly modular design as well as by convenient configuration through MS-Excel tables. |
| Paper |
Tecan Application Note (not peer reviewed):
Grünberg R, van der Sloot A., Xu X., Tyers M. (2015) Fully
integrated plating and colony picking for synthetic
biology workflows.
[PDF]
[PDF via Tecan]
(We are working on a proper paper. The Tecan folks also show-cased our platform in their Tecan Journal) |
 |
postdoc / research associate (2010-2013) |
|---|---|
| hiFRET -- the Power of Weak Interactions | |
|
Center for Genomic Regulation (CRG), Barcelona IRIC, University of Montreal |
|
| Poster | |
| Abstract |
FRET (Fluorescent Resonance Energy Transfer) is currently the only
practical method to localize and image protein interactions in live
cells. But FRET sensor design is complicated by the need to bring the
two fluorescent proteins (donor and acceptor) very close together. To
overcome this limitation, we engineered very weak affinities between
donor and acceptor probes. Not unlike the magnetic cover of a tablet
computer, once donor and acceptor are brought together by the
interaction of interest, our helper affinity is pulling them into a
better orientation for strong FRET.
We realized these helper interactions in two different ways. First, we created an electrostatic "encounter complex" between FRET probes using computational design. In a second approach we instead attached specific but very weak "protein interaction modules" from natural signalling proteins. This second variant can be immediately applied to any of a wide variety of modern fluorescent proteins. We tested hiFRET probes for the detection of different protein interactions in vitro and in live cells. Thanks to hiFRET, we were the first to image the interaction between BRaf and CRaf proteins in response to a new anti-cancer drug. This interaction offers tumors a way to escape the cancer treatment and is therefore also of medical interest. |
| Paper |
Grünberg R, Burnier JV, Ferrar T,
Beltran-Sastre V, Stricher F, van der
Sloot A, Garcia-Olivas R, Mallabiabarrena A,
Sanjuan X, Zimmermann T, Serrano L (2013):
Engineering of weak helper interactions for high-efficiency FRET probes.
Nature Methods. News and Views by Kees Jalink. (requires subsription) |
 |
postdoc (2006-2010) |
|---|---|
| Protein Synthetic Biology | |
| Center for Genomic Regulation (CRG), Barcelona | |
| Poster | HFSP Fellows meeting, Tokyo, June 2009 |
| Abstract | Can synthetic proteins and sophisticated protein circuits be built from standard parts? Can we decompose dynamic protein networks into re-usable devices? What are the best strategies and methods for protein systems engineering? These are some of the questions that I have been working on since switching back to the wet lab bench. Despite all their complexity (see my previous projects below), I think that proteins are, in fact, a very good material for systems engineering. A focus on modular protein-protein interactions may be our best starting point for the manipulation of natural and the design of synthetic protein networks. In a proof-of-concept study, we have constructed 25 synthetic two-domain proteins from BioBrick-formatted standardized "parts", purified many of them and characterized different combinations of interaction input and output "devices" biophysically (using FRET and surface plasmon resonance). Our parts are available for free re-use from the Registry of Standard Biological Parts and a list of protocols is maintained on OpenWetware. |
| Paper |
Grünberg R, Ferrar T, van der Sloot A, Constante M, Serrano L (2010)
Building blocks for protein interaction devices. Nucleic Acids Research
doi: 10.1093/nar/gkq152.
[Full text] Grünberg R, Serrano L (2010): Strategies for Protein Synthetic Biology. (review) Nucleic Acids Research in press. |
 |
postdoc (2007-2009) |
|---|---|
| (Voronoi-) Shelling of Protein Interfaces | |
| Collaboration with Frederic Cazals (INRIA Sophia-Antipolis), Benjamin Bouvier (CNRS Lyon) and Michael Nilges (Institut Pasteur) | |
| Abstract | Protein-protein interfaces are, traditionally, defined by different "ad-hoc" criteria such as atom contacts or changes in surface areas. However, these descriptions are rather arbitrary and complicate the systematic comparison of protein interface architectures. We here extend a fast, parameter-free and purely geometric definition of protein interfaces and introduce the shelling order of Voronoi facets (VSO) as a novel measure for an atom's depth inside the interface. VSO can predict the solvent exchange in different interface regions, that is, the occurence of "dry" or "wet" residues. In fact, the seemingly complex water dynamics at protein interfaces appears largely controlled by geometry. Also the sequence conservation and residue composition of interfaces is often "radially" structured. Voronoi Shelling Order thus reveals clear geometric patterns in protein interface composition, function and dynamics and facilitates the comparative analysis of protein-protein interactions. |
| Paper |
Bouvier B*, Grünberg R*, Nilges M, Cazals F (2009):
Shelling the Voronoi interface of protein-protein complexes
reveals patterns of residue conservation, dynamics, and composition.
Proteins 15;76(3):677-92.
[Abstract] (* shared first co-authors) |
 |
PhD project (2001 - 2005) |
|---|---|
| The dynamics of protein-protein binding | |
| Institut Pasteur, Paris | |
| Talk | Protein flexibility and entropy on the edge of binding (International Biophysics Congress, August 2005, Montpellier) |
| Poster | Conference on "Modeling of Protein Interactions in Genomes", June 2003, Stony Brook, New York; June 2005, Kansas |
| Abstract |
The interplay between protein-protein binding and structural dynamics
is poorly understood. We selected a set of 17 protein-protein
complexes for which the three-dimensional structures of both free
components and the complex are available. Based on the analysis of
molecular dynamics simulations in explicit water for each of these 51
systems, we find: (1) most uncomplexed binding sites are more flexible
than the remaining surface; (2) as expected, they loose conformational
freedom upon complex formation; (3) however, in the majority of cases,
binding does not restrict the overall motion of proteins. We
calculated the change in conformational entropy from longer
simulations on 7 complexes (21 systems) using a new method based on
quasiharmonic analysis. Two small complexes and an antibody-antigen
system exhibited a significant loss whereas three larger complexes
showed increased or unchanged conformational entropy. Molecular fluctuations excert important influence both on the speed of recognition and the stability of protein complexes. Structure dynamics may be the missing link in our understanding of this fundamental process. This is the largest study so far on the influence of complex formation on protein flexibility and conformational entropy. |
| Paper |
Grünberg, R., M. Nilges, J. Leckner (2006):
Flexibility and entropy in protein-protein
binding. Structure 14(4), 683-93
[PDF] |
 |
PhD project (2001 - 2004) |
|---|---|
| A model of flexible protein-protein recognition | |
| Institut Pasteur, Paris | |
| Talk |
Complementarity of structure ensembles in protein-protein
binding
(CECAM workshop on "Flexible Macromolecular Docking", April 2004, Lyon) |
| Abstract |
Protein-protein association is often accompanied by changes in
receptor and ligand structure. This interplay between protein
flexibility and protein-protein recognition is currently the
largest obstacle to our understanding and also to the reliable
prediction of protein complexes. We performed two sets of
molecular dynamics simulations for the unbound receptor and
ligand structures of 17 protein complexes and applied
shape-driven rigid body docking to all combinations of
representative snapshots. The cross docking of structure
ensembles increased the chances to find near native
solutions. The free ensembles appeared to contain multiple
complementary conformations. These were in general not related
to the bound structure. We suggest that protein-protein
binding follows a three-step mechanism of diffusion, free
conformer selection and folding. This model combines
previously conflicting ideas and is in better agreement with
the current data on interaction forces, time scales, and
kinetics.
|
| Paper |
*Grünberg, R., *J. Leckner, M. Nilges (2004): Complementarity of
structure ensembles in protein-protein
binding. Structure 12, 2125-2136. [PubMed]
[pre-print] (with minor differences)
(* shared first co-authors)
Comment by M. Eisenstein in the same issue: Introducing a 4th dimension to protein-protein docking. [PDF] Quote of our model in a commentary by Boehr and Wright in Science: How do proteins interact? |
 |
PhD project (2002 - 2004) |
|---|---|
| CROW - A data integration platform for the semantic web | |
| Institut Pasteur, Paris | |
| Talk | Crowsing the semantic web (Journee bioinformatique, February 2004, Institut Pasteur) |
| Poster | Pacific Symposium on Biocomputing, January 2004, Hawaii |
| Abstract |
Crow (Computational Representation Of Whatever) is a
(alpha-stage) Java library for an anarchist approach
to data integration (i.e. link whatever you want
whenever you want to whereever you want). The library
is tailored (but not restricted) to the work with
semantic web documents. It allows the mining and
manipulation of distributed, heterogeneous,
chaotically cross-linked data. The design is
multi-threaded, event-based, and supports plugins and
third-party data types. It containes a simple Python
(Jython) interface for interactive
work.
The library is freely available under the GPL.
We currently use it to select targets for
protein-protein docking from sets of predicted protein interactions
and various other information.
SourceForge project... |
| Paper | I somehow never got to publish this...but the code is on sourceforge and well documented. If anybody wants to join me converting this into a Python library... send me an e-mail ;-) |
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PhD project (warm up) (06/2000 - 12/2001) |
|---|---|
| The dynamic topology of spectrin repeats | |
| EMBL, Heidelberg and Institut Pasteur, Paris | |
| Talk | Stretching a molecular shock absorber
(Congress des jeunes chercheurs, 18/06/2003, Institut Pasteur) |
| Poster | Gordon conference on "Protein folding dynamics", January 2002, Ventura, U.S. |
| Abstract |
Spectrin repeats are triple-helical domains found in many
proteins that are regularly subjected to mechanical stress. My collaborators
studied the behavior of a wild-type spectrin repeat and
two mutants with atomic force microscopy. I interpreted the results
of these single molecule experiments with steered molecular dynamics
simulations. The experiments indicated that spectrin repeats can form
stable unfolding intermediates when subjected to external forces. In
the simulations the unfolding proceeded via a variety of
pathways. Stable intermediates were associated to kinking of the
central helix close to a proline residue. A mutant stabilizing the
central helix showed no intermediates in experiments, in agreement
with the simulations. Simulations and experiments hence suggest a "hyphenation
point" within the domain and hint at the existence of force-resistant
intermediates with non-native topology. The simulations also explained the
broad variance of experimental unfolding lenghts. The combination of both
effects would allow for a smooth and elastic response to external forces
over a wide range of extensions. Spectrin molecules seem to be optimized for
elasticity on all scales of their molecular architecture.
|
| Paper | *Altmann S.M., R. *Grünberg, P.F. Lenne, J. Ylänne, A. Raae, K. Herbert, M. Saraste, M. Nilges, J.K. Hörber (2002): Pathways and intermediates in forced unfolding of spectrin repeats. Structure 10, 1085-1096. [PubMed] (* shared first co-authors) |
 |
Diploma Project (03/99 - 10/99) |
|---|---|
| Rational Design of Trypsin Variants for Peptide Synthesis | |
| University of Leipzig, under the guidance of Dr. Frank Bordusa | |
| Talk | Hacking Trypsin (given on June 25st 1999 in Leipzig) |
| Abstract |
Based on their catalytic mechanism it is possible to employ serine proteases like
trypsin for enzymatic peptide bond formation. Especially, substrate mimetics -
a new class of artificial substrates, allow for efficient ligations of amino acid
derivatives and peptides, as Bordusa et al. demonstrated in recent years.
However, if product or starting material contain specific cleavage sites recognised
by the protease, this ligation is counteracted by the (natural) proteolytic reaction.
A promising approach to overcome this problem is the direct manipulation of the enzyme's natural specifity. I prepared and isolated two variants of rat-trypsin with altered S1-binding pocket (D189A and D189A/S190A). The proteins were examined by HPLC-based acyl-transfer experiments revealing promising catalytic properties as well as an impaired specifity towards natural trypsin and chymotrypsin substrates. Parallely, I employed the program AutoDock to simulate the binding of several substrates to the altered enzymes in silico. With these docking studies I could elucidate, in part unexpected, experimental results on a structural basis. |
| Paper | Grünberg, R., I. Domgall, R. Günther, H.-J. Hofmann, and F. Bordusa (2000): Peptide Bond Formation Mediated by Substrate Mimetics: Structure-guided Optimisation of Trypsin for Synthesis. Eur J Biochem 267(24), 7024-30. |
|
4th-year-project (02/98 - 06/98) |
|---|---|
| Analysing the adc promoter in Clostridium beijerinckii with the gusA reporter gene |
|
| University of Wales, Aberystwyth, under the guidance of Dr. Adriana Ravagnani and Prof. Michael Young | |
| Talk | A reporter gene for Chlostridium (given in April 1998 in Aberystwyth) |
| Abstract | I introduced ß-D-glucuronidase (gusA) of Escherichia
coli as reporter gene in Clostridium beijerinckii, so far lacking a
versatile reporter system, by constructing
the promoter probe vector pRGus. This vector was
employed to evaluate the significance of three putative "OA box" sequence motifs within
the promoter of the acetoacetate decarboxylase gene (adc) in vivo.
I fused the adc promoter, as well as a variant with destroyed OA boxes,
to the gusA gene in pRGus and introduced them into C. beijerinckii
by electroporation. Preliminary tests revealed a high ß-D-glucuronidase (GUS) activity in transformants containing the wild type adc promoter compared with no detectable GUS activity in case of the construct with destroyed OA boxes. Since OA boxes are recognition sequences for the Spo0A transcription factor, these findings indicate that the expression of the adc gene is directly regulated by Spo0A. Acetoacetate decarboxylase is involved in the metabolic conversion of starches (and various other substrates) into commercially significant solvents like acetone and butanol. The gusA reporter gene will facilitate further investigation into the trancsiptional regulation of this pathway. |
| Paper | Ravagnani, A., K. C. B. Jennert, E. Steiner, R. Grünberg, J. R. Jefferies, S. R. Wilkinson, D. I. Young, E. C. Tidswell, D. P. Brown, P. Youngman, J. G. Morris & M. Young. (2000): Spo0A directly controls the switch from acid to solvent production in solvent-forming clostridia. Molecular Microbiology 37(5), 1172-85. |
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