Activity
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Yesterday, we celebrated 25 years of the Bornscheuer group (https://lnkd.in/dVYWaYU7) at the Universität Greifswald and enjoyed nice lectures given…
Yesterday, we celebrated 25 years of the Bornscheuer group (https://lnkd.in/dVYWaYU7) at the Universität Greifswald and enjoyed nice lectures given…
Liked by Alberto Nobili
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I am personally very happy and proud about the news that the trial we designed some years ago as pivotal Phase 3 trial inMIND in R/R FL for the CD19…
I am personally very happy and proud about the news that the trial we designed some years ago as pivotal Phase 3 trial inMIND in R/R FL for the CD19…
Liked by Alberto Nobili
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Every chapter has an ending. Earlier in August, I resigned from my role as Head of Bioengineering at ODDITY. Beyond drugs, I’ve always believed in…
Every chapter has an ending. Earlier in August, I resigned from my role as Head of Bioengineering at ODDITY. Beyond drugs, I’ve always believed in…
Liked by Alberto Nobili
Experience
Education
Licenses & Certifications
Volunteer Experience
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Defensive Coach Assistant
RAMS Milano American Football Team a.s.d.
- 1 year 1 month
Social Services
I supported the team's head coach during the preparation and development of the regular training and on the gameday. My main goal was to be useful to the team and learn anything possible from the experience.
Publications
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Light-Controllable Binary Switch Activation of CAR T Cells
ChemMedChem
Major challenges to chimeric antigen receptor (CAR) T cell therapies include uncontrolled immune activity, off-tumor toxicities and tumor heterogeneity. To overcome these challenges, we engineered CARs directed against small molecules. By conjugating the same small molecule to distinct tumor targeting antibodies, we show that small molecule specific-CAR T cells can be redirected to different tumor antigens. Such binary switches allow control over the degree of CAR T cell activity and enables…
Major challenges to chimeric antigen receptor (CAR) T cell therapies include uncontrolled immune activity, off-tumor toxicities and tumor heterogeneity. To overcome these challenges, we engineered CARs directed against small molecules. By conjugating the same small molecule to distinct tumor targeting antibodies, we show that small molecule specific-CAR T cells can be redirected to different tumor antigens. Such binary switches allow control over the degree of CAR T cell activity and enables simultaneous targeting of multiple tumor associated antigens. We also demonstrate that ultraviolet light sensitive caging of small molecules blocks CAR T cell activation. Exposure to ultraviolet light, uncaged small molecules and restored CAR T cell-mediated killing. Together, our data demonstrate that a light-sensitive caging system enables an additional level of control over tumor cell killing, which could improve the therapeutic index of CAR T cell therapies.
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Creation of (R)-Amine Transaminase Activity within an α-Amino Acid Transaminase Scaffold
ACS Chem. Biol.
The enzymatic transamination of ketones into (R)-amines represents an important route for accessing a range of pharmaceuticals or building blocks. Although many publications have dealt with enzyme discovery, protein engineering, and the application of (R)-selective amine transaminases [(R)-ATA] in biocatalysis, little is known about the actual in vivo role and how these enzymes have evolved from the ubiquitous α-amino acid transaminases (α-AATs). Here, we show the successful introduction of an…
The enzymatic transamination of ketones into (R)-amines represents an important route for accessing a range of pharmaceuticals or building blocks. Although many publications have dealt with enzyme discovery, protein engineering, and the application of (R)-selective amine transaminases [(R)-ATA] in biocatalysis, little is known about the actual in vivo role and how these enzymes have evolved from the ubiquitous α-amino acid transaminases (α-AATs). Here, we show the successful introduction of an (R)-transaminase activity in an α-amino acid aminotransferase with one to six amino acid substitutions in the enzyme’s active site. Bioinformatic analysis combined with computational redesign of the d-amino acid aminotransferase (DATA) led to the identification of a sextuple variant having a specific activity of 326 milliunits mg–1 in the conversion of (R)-phenylethylamine and pyruvate to acetophenone and d-alanine. This value is similar to those of natural (R)-ATAs, which typically are in the range of 250 milliunits mg–1. These results demonstrate that (R)-ATAs can evolve from α-AAT as shown here for the DATA scaffold.
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CorNet: Assigning function to networks of co-evolving residues by automated literature mining
PLoS One
CorNet is a web-based tool for the analysis of co-evolving residue positions in protein super-family sequence alignments. CorNet projects external information such as mutation data extracted from literature on interactively displayed groups of co-evolving residue positions to shed light on the functions associated with these groups and the residues in them. We used CorNet to analyse six enzyme super-families and found that groups of strongly co-evolving residues tend to consist of residues…
CorNet is a web-based tool for the analysis of co-evolving residue positions in protein super-family sequence alignments. CorNet projects external information such as mutation data extracted from literature on interactively displayed groups of co-evolving residue positions to shed light on the functions associated with these groups and the residues in them. We used CorNet to analyse six enzyme super-families and found that groups of strongly co-evolving residues tend to consist of residues involved in a same function such as activity, specificity, co-factor binding, or enantioselectivity. This finding allows to assign a function to residues for which no data is available yet in the literature. A mutant library was designed to mutate residues observed in a group of co-evolving residues predicted to be involved in enantioselectivity, but for which no literature data is available yet. The resulting set of mutations indeed showed many instances of increased enantioselectivity.
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NewProt - a protein engineering portal
Protein Engineering Design and Selection
The NewProt protein engineering portal is a one-stop-shop for in silico protein engineering. It gives access to a large number of servers that compute a wide variety of protein structure characteristics supporting work on the modification of proteins through the introduction of (multiple) point mutations. The results can be inspected through multiple visualizers. The HOPE software is included to indicate mutations with possible undesired side effects. The Hotspot Wizard software is embedded for…
The NewProt protein engineering portal is a one-stop-shop for in silico protein engineering. It gives access to a large number of servers that compute a wide variety of protein structure characteristics supporting work on the modification of proteins through the introduction of (multiple) point mutations. The results can be inspected through multiple visualizers. The HOPE software is included to indicate mutations with possible undesired side effects. The Hotspot Wizard software is embedded for the design of mutations that modify a proteins' activity, specificity, or stability. The NewProt portal is freely accessible at http://newprot.cmbi.umcn.nl/ and http://newprot.fluidops.net/
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Bioinformatic analysis of fold-type III PLP-dependent enzymes discovers multimeric racemases
Applied Microbiology and Biotechnology
Pyridoxal-5′-phosphate (PLP)-dependent enzymes are ubiquitous in nature and catalyze a variety of important metabolic reactions. The fold-type III PLP-dependent enzyme family is primarily comprised of decarboxylases and alanine racemases. In the development of a multiple structural alignment database (3DM) for the enzyme family, a large subset of 5666 uncharacterized proteins with high structural, but low sequence similarity to alanine racemase and decarboxylases was found. Compared to these…
Pyridoxal-5′-phosphate (PLP)-dependent enzymes are ubiquitous in nature and catalyze a variety of important metabolic reactions. The fold-type III PLP-dependent enzyme family is primarily comprised of decarboxylases and alanine racemases. In the development of a multiple structural alignment database (3DM) for the enzyme family, a large subset of 5666 uncharacterized proteins with high structural, but low sequence similarity to alanine racemase and decarboxylases was found. Compared to these two classes of enzymes, the protein sequences being the object of this study completely lack the C-terminal domain, which has been reported important for the formation of the dimer interface in other fold-type III enzymes. The 5666 sequences cluster around four protein templates, which also share little sequence identity to each other. In this work, these four template proteins were solubly expressed in Escherichia coli, purified, and their substrate profiles were evaluated by HPLC analysis for racemase activity using a broader range of amino acids. They were found active only against alanine or serine, where they exhibited Michaelis constants within the range of typical bacterial alanine racemases, but with significantly lower turnover numbers. As the already described racemases were proposed to be active and appeared to be monomers as judged from their crystal structures, we also investigated this aspect for the four new enzymes. Here, size exclusion chromatography indicated the presence of oligomeric states of the enzymes and a native-PAGE in-gel assay showed that the racemase activity was present only in an oligomeric state but not as monomer. This suggests the likelihood of a different behavior of these enzymes in solution compared to the one observed in crystalline form.
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Two Subtle Amino Acid Changes in a Transaminase Substantially Enhance or Invert Enantiopreference in Cascade Syntheses
ChemBioChem
Amine transaminases (ATAs) are powerful enzymes for the stereospecific production of chiral amines. However, the synthesis of amines incorporating more than one stereocenter is still a challenge. We developed a cascade synthesis to access optically active 3-alkyl-substituted chiral amines by combining two asymmetric synthesis steps catalyzed by an enoate reductase and ATAs. The ATA wild type from Vibrio fluvialis showed only modest enantioselectivity (14 % de) in the amination of…
Amine transaminases (ATAs) are powerful enzymes for the stereospecific production of chiral amines. However, the synthesis of amines incorporating more than one stereocenter is still a challenge. We developed a cascade synthesis to access optically active 3-alkyl-substituted chiral amines by combining two asymmetric synthesis steps catalyzed by an enoate reductase and ATAs. The ATA wild type from Vibrio fluvialis showed only modest enantioselectivity (14 % de) in the amination of (S)-3-methylcyclohexanone, the product of the enoate-reductase-catalyzed reaction step. However, by protein engineering we created two variants with substantially improved diastereoselectivities: variant Leu56Val exhibited a higher R selectivity (66 % de) whereas the Leu56Ile substitution caused a switch in enantiopreference to furnish the S-configured diastereomer (70 % de). Addition of 30 % DMSO further improved the selectivity and facilitated the synthesis of (1R,3S)-1-amino-3-methylcyclohexane with 89 % de at 87 % conversion.
Other authorsSee publication -
Simultaneous Use of in Silico Design and a Correlated Mutation Network as a Tool To Efficiently Guide Enzyme Engineering
ChemBioChem 2015, n/a, n/a
In order to improve the efficiency of directed evolution experiments, in silico multiple-substrate clustering was combined with an analysis of the variability of natural enzymes within a protein superfamily. This was applied to a Pseudomonas fluorescens esterase (PFE I) targeting the enantioselective hydrolysis of 3-phenylbutyric acid esters. Data reported in the literature for nine substrates were used for the clustering meta-analysis of the docking conformations in wild-type PFE I, and this…
In order to improve the efficiency of directed evolution experiments, in silico multiple-substrate clustering was combined with an analysis of the variability of natural enzymes within a protein superfamily. This was applied to a Pseudomonas fluorescens esterase (PFE I) targeting the enantioselective hydrolysis of 3-phenylbutyric acid esters. Data reported in the literature for nine substrates were used for the clustering meta-analysis of the docking conformations in wild-type PFE I, and this highlighted a tryptophan residue (W28) as an interesting target. Exploration of the most frequently, naturally occurring amino acids at this position suggested that the reduced flexibility observed in the case of the W28F variant leads to enhancement of the enantioselectivity. This mutant was subsequently combined with mutations identified in a library based on analysis of a correlated mutation network. By interrogation of <80 variants a mutant with 15-fold improved enantioselectivity was found.
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Engineering the Active Site of the Amine Transaminase from Vibrio fluvialis for the Asymmetric Synthesis of Aryl–Alkyl Amines and Amino Alcohols
ChemCatChem 2015, n/a-n/a
The constrained active site of the amine-transaminase from Vibrio fluvialis was engineered in order to achieve the conversion of bulky ketones. This led to the discovery of mutants with a 30-fold increase in activity which enabled the asymmetric synthesis of (S)-phenylbutylamine and (R)-phenylglycinol
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Bioinformatic analysis of a PLP-dependent enzyme superfamily suitable for biocatalytic applications
Biotechnol. Adv. 2015 n/a-n/a
In this review we analyse structure/sequence–function relationships for the superfamily of PLP-dependent enzymes with special emphasis on class III transaminases. Amine transaminases are highly important for applications in biocatalysis in the synthesis of chiral amines. In addition, other enzyme activities such as racemases or decarboxylases are also discussed. The substrate scope and the ability to accept chemically different types of substrates are shown to be reflected in conserved patterns…
In this review we analyse structure/sequence–function relationships for the superfamily of PLP-dependent enzymes with special emphasis on class III transaminases. Amine transaminases are highly important for applications in biocatalysis in the synthesis of chiral amines. In addition, other enzyme activities such as racemases or decarboxylases are also discussed. The substrate scope and the ability to accept chemically different types of substrates are shown to be reflected in conserved patterns of amino acids around the active site. These findings are condensed in a sequence–function matrix, which facilitates annotation and identification of biocatalytically relevant enzymes and protein engineering thereof.
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Improved thermostability of a Bacillus subtilis esterase by domain exchange
Appl. Microbiol. Biotechnol. 2014, 98, 1719-172
A moderately thermostable esterase from Geobacillus stearothermophilus (BsteE) and its homolog from Bacillus subtilis (BsubE) show a high structural similarity with more than 95% homology and 74% amino acid identity. Interestingly, their thermal stability differs significantly by 30 °C in their melting temperature. In order to identify the positions that are responsible for this difference, most of the flexible amino acids assumed to confer instability were found to be in the cap region. For…
A moderately thermostable esterase from Geobacillus stearothermophilus (BsteE) and its homolog from Bacillus subtilis (BsubE) show a high structural similarity with more than 95% homology and 74% amino acid identity. Interestingly, their thermal stability differs significantly by 30 °C in their melting temperature. In order to identify the positions that are responsible for this difference, most of the flexible amino acids assumed to confer instability were found to be in the cap region. For this reason, a 30 amino acid long cap domain fragment containing ten differing positions derived from BsteE was incorporated into the homologous gene encoding for the more labile BsubE by spliced overlap-extension PCR. The melting temperature of the two wild-type esterases and the mutant was evaluated by circular dichroism spectroscopy, while the kinetic parameters and the stability were determined with a photometric assay. The cap domain mutant maintained its activity, with a catalytic efficiency more similar to BsteE, while it exhibited an increase of the melting temperature by 4 °C compared to BsubE. Additional point mutations based on the differences of the parent enzymes gave a further increase of the thermostability up to 11 °C compared to BsubE; however, a significant reduction in activity was observed.
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Use of 'small but smart' libraries to enhance the enantioselectivity of an esterase from Bacillus stearothermophilus towards tetrahydrofuran-3-yl acetate
FEBS J. 2013, 280, 3084-3093
Two libraries of simultaneous double mutations in the active site region of an esterase from Bacillus stearothermophilus were constructed to improve the enantioselectivity in the hydrolysis of tetrahydrofuran-3-yl acetate. As screening of large mutant libraries is hampered by the necessity for GC/MS analysis, mutant libraries were designed according to a 'small but smart' concept. The design of focused libraries was based on data derived from a structural alignment of 3317 amino acid sequences…
Two libraries of simultaneous double mutations in the active site region of an esterase from Bacillus stearothermophilus were constructed to improve the enantioselectivity in the hydrolysis of tetrahydrofuran-3-yl acetate. As screening of large mutant libraries is hampered by the necessity for GC/MS analysis, mutant libraries were designed according to a 'small but smart' concept. The design of focused libraries was based on data derived from a structural alignment of 3317 amino acid sequences of α/β-hydrolase fold enzymes with the bioinformatic tool 3DM. In this way, the number of mutants to be screened was substantially reduced as compared with a standard site-saturation mutagenesis approach. Whereas the wild-type esterase showed only poor enantioselectivity (E = 4.3) in the hydrolysis of (S)-tetrahydrofuran-3-yl acetate, the best variants obtained with this approach showed increased E-values of up to 10.4. Furthermore, some variants with inverted enantiopreference were found.
Honors & Awards
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Marie Curie Fellow
ENEFP European Network on Directed Evolution of Functional Proteins
Languages
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Italian
Native or bilingual proficiency
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Spanish
Native or bilingual proficiency
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English
Full professional proficiency
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German
Professional working proficiency
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Catalan
Elementary proficiency
More activity by Alberto
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Congratulations to this year's cohort, the third and largest of the Innovation Fellows Program, led by aMoon General Partner, Dr Yaron Daniely. We…
Congratulations to this year's cohort, the third and largest of the Innovation Fellows Program, led by aMoon General Partner, Dr Yaron Daniely. We…
Liked by Alberto Nobili
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was privileged to have my commentary be published in Nature's World View last Th. https://lnkd.in/eDK3UCw9
was privileged to have my commentary be published in Nature's World View last Th. https://lnkd.in/eDK3UCw9
Liked by Alberto Nobili
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Great to see our friends from WhiteLab Genomics leading the way! #Bayer #CoLab David Del Bourgo Nathan Lee
Great to see our friends from WhiteLab Genomics leading the way! #Bayer #CoLab David Del Bourgo Nathan Lee
Shared by Alberto Nobili
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Roof Deck Party! Join us for fun and games at Portal Playground on Sept 16th at Bayer Co.Lab Cambridge. Register: https://lu.ma/gs7ptchr
Roof Deck Party! Join us for fun and games at Portal Playground on Sept 16th at Bayer Co.Lab Cambridge. Register: https://lu.ma/gs7ptchr
Liked by Alberto Nobili
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An outstanding 5 years of Nucleate. Check out our first Impact Report! https://lnkd.in/gD7NPP3K I’m proud of the work we’ve accomplished, and…
An outstanding 5 years of Nucleate. Check out our first Impact Report! https://lnkd.in/gD7NPP3K I’m proud of the work we’ve accomplished, and…
Liked by Alberto Nobili
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This insight into the impact of myelin on physical performance is fascinating! A compelling reminder of the importance of staying active. Next time I…
This insight into the impact of myelin on physical performance is fascinating! A compelling reminder of the importance of staying active. Next time I…
Liked by Alberto Nobili
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Nature Magazine just recently published an article written by Melanie Senior that sheds light on the latest trends in #VC #financing within…
Nature Magazine just recently published an article written by Melanie Senior that sheds light on the latest trends in #VC #financing within…
Liked by Alberto Nobili
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Things continue to advance here at Bayer Co.Lab. Hot on the heels of our 50-day mark for Bayer Co.Lab Shanghai, we’re now launching the 100-day…
Things continue to advance here at Bayer Co.Lab. Hot on the heels of our 50-day mark for Bayer Co.Lab Shanghai, we’re now launching the 100-day…
Liked by Alberto Nobili
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The #Lamborghini #Temerario – unveiled today at the Monterey Car Week - represents the second and incredibly important milestone for Sonus faber…
The #Lamborghini #Temerario – unveiled today at the Monterey Car Week - represents the second and incredibly important milestone for Sonus faber…
Liked by Alberto Nobili
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First Fierce, now Stat… the “price controls are actually fine for innovation” argument sure seems irresistibly contagious. In this case, STAT…
First Fierce, now Stat… the “price controls are actually fine for innovation” argument sure seems irresistibly contagious. In this case, STAT…
Liked by Alberto Nobili
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Introducing Lamborghini Temerario: the Sound of Italian Excellence.🌟 Unveiled today at Monterey Car Week, the new Temerario is the most advanced…
Introducing Lamborghini Temerario: the Sound of Italian Excellence.🌟 Unveiled today at Monterey Car Week, the new Temerario is the most advanced…
Liked by Alberto Nobili
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Temerario, the last Lamborghini project I worked on, has been unveiled - and what better stage than my adopted home, California! Witnessing this…
Temerario, the last Lamborghini project I worked on, has been unveiled - and what better stage than my adopted home, California! Witnessing this…
Liked by Alberto Nobili
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