Roberto Chica Lab

Announcement for the 6th Protein Engineering Canada Conference, to be held June 22nd-24th in Ottawa, Canada. The image shows a protein structure in front of a picture of Parliament Hill and Chateau Laurier in Ottawa.

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Save the date!

The 6th Protein Engineering Canada Conference will be held on June 22-24 in Ottawa, Canada.

Abstract submission and registration are open!

More information here: event.fourwaves.com/pec2026/pages

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Speakers of the 6th Protein Engineering Canada Conference include:

Bill DeGrado
Joelle Pelletier
Tim Whitehead
Anastassia Vorobieva
Lucy Colwell
Ai Niitsu
@joannas.bsky.social
@nickpolizzi.bsky.social
@possuhuanglab.bsky.social
@paolalaurino.bsky.social
@stephanhammer.bsky.social
and more!

Announcement for the 6th Protein Engineering Canada Conference, to be held June 22nd-24th in Ottawa, Canada. The image shows a protein structure in front of a picture of Parliament Hill and Chateau Laurier in Ottawa.

1/
Save the date!

The 6th Protein Engineering Canada Conference will be held on June 22-24 in Ottawa, Canada.

Abstract submission and registration are open!

More information here: event.fourwaves.com/pec2026/pages

Distal mutations enhance catalysis in designed enzymes by facilitating substrate binding and product release - Nature Communications Distal mutations, though far from the active site, enhance Kemp eliminase catalysis by tuning conformational dynamics that facilitate substrate binding and product release, thereby promoting the full catalytic cycle.

Thrilled to share that our latest article is now out in final form! A great collaboration with @fraserlab.com and @silviaosuna.bsky.social.

Distal mutations enhance catalysis in designed enzymes by facilitating substrate binding and product release

www.nature.com/articles/s41...

Distal Mutations in a Designed Retro-Aldolase Alter Loop Dynamics to Shift and Accelerate the Rate-Limiting Step Amino acid residues distant from an enzyme’s active site are known to influence catalysis, but their mechanistic contributions to the catalytic cycle remain poorly understood. Here, we investigate the...

Our latest article is now published online! In collaboration with @thompson-lab.bsky.social, Marc Garcia-Borràs, and @ferranfeixas.bsky.social.

Distal Mutations in a Designed Retro-Aldolase Alter Loop Dynamics to Shift and Accelerate the Rate-Limiting Step

pubs.acs.org/doi/full/10....

Enzyme-like proteins by computational design | PNAS We report the development and initial experimental validation of a computational design procedure aimed at generating enzyme-like protein catalys...

Similarly, enzyme function can be designed de novo by creating a new active site within a natural protein scaffold that lacks the target activity, even if that catalytic function exists in nature.

See below for an early example:

www.pnas.org/doi/full/10....

De novo design of a four-fold symmetric TIM-barrel protein with atomic-level accuracy - Nature Chemical Biology Despite substantial effort, the de novo design of a stable TIM-barrel protein fold has remained elusive. A Rosetta-based computational strategy identifies a unique 184-residue sequence that adopts a T...

No, I don’t think that’s necessarily implied. For example, a TIM barrel can be designed from scratch without referencing any specific natural sequence or structure, even if this fold exists in nature. I consider this de novo design. See example below:

www.nature.com/articles/nch...

Characterization of a Helical Protein Designed from First Principles The question of how the primary amino acid sequence of a protein determines its three-dimensional structure is still unanswered. One approach to this problem involves the de novo design of model pepti...

The design and creation of a protein sequence, structure or function from scratch, rather than modifying a pre-existing sequence, structure or function.

An early pioneer of this field is Bill DeGrado, see below.

www.science.org/doi/10.1126/...

Overall, our study:
✅ Introduces a new strategy to transform minimal protein scaffolds into biocatalysts
✅ Provides mechanistic insights from crystallography & molecular dynamics
✅ Opens the door to designing custom lids for more complex reactions, which we’re now exploring

Thanks for reading! 🧵🧬

The crystal structure (blue) aligns closely with the design model (minimal TIM barrel and lid colored white and magenta, respectively).

Our crystal structure validated the designed fold, confirming that the lid was correctly folded.

However, a subtle 1.8 Å lid shift disrupted a key catalytic contact, likely contributing to the modest activity. But structural analysis reveals paths to improve activity in the next round of design!

Michaelis-Menten plot of KempTIM4 showing saturation kinetics.

One of our designs, KempTIM4, showed catalytic efficiency comparable to many first-round de novo Kemp eliminases generated by traditional methods.

Building a custom lid onto a minimal, de novo TIM barrel using CANVAS.

Using CANVAS, we designed a structural lid onto a minimal, de novo TIM barrel to anchor catalytic residues and create an active site for the Kemp elimination reaction.

De novo enzyme design using CANVAS.

TIM barrels are among nature’s most powerful enzyme scaffolds but making them from scratch with catalytic function has been a challenge.

Enter CANVAS: a computational pipeline combining Triad, RFdiffusion & ProteinMPNN to customize minimal TIM barrels into functional enzymes.

Customizing the Structure of a Minimal TIM Barrel to Craft a De Novo Enzyme The TIM barrel is the most prevalent fold in natural enzymes, supporting efficient catalysis of diverse chemical reactions. While de novo TIM barrels have been successfully designed, their minimalisti...

In collaboration with @birtehoecker.bsky.social, we’ve unlocked enzymatic activity in a minimal de novo TIM barrel by designing a custom lid for catalysis! 🧵👇
#ProteinDesign #EnzymeDesign

Customizing the Structure of a Minimal TIM Barrel to Create a De Novo Enzyme
www.biorxiv.org/content/10.1...

Congratulations! Looking forward to seeing all the exciting science that will come out of your lab! 🧪

Research Position (PhD) in organic chemistry and b... <div style="text-align: justify;">The&nbsp; research&nbsp; group&nbsp; „Organic&nbsp; Chemistry&nbsp; and&nb...

Join us! We are looking for a new team member (PhD student) with strong background in organic chemistry.
🙏 RETWEET (We want to recruit internationally)

Organic chemistry meets #DirectedEvolution
Highly interdisciplinary & passionate research group

uni-bielefeld.hr4you.org/job/view/433...

Non-canonical amino acid from PDB ID 8W3Z shown chelating to a magnesium ion. Image made with PyMol. https://www.rcsb.org/structure/8W3Z https://www.pymol.org

Protein Engineering, Design & Selection (PEDS) invites contributions to a Special Collection titled, “Non-Canonical Amino Acids", with guest editors Prof. Huiwang Ai (Virginia) and Prof. Peng Chen (Peking). Send us your best work!
academic.oup.com/peds/pages/c...

Zero-shot design of drug-binding proteins via neural selection-expansion Computational design of molecular recognition remains challenging despite advances in deep learning. The design of proteins that bind to small molecules has been particularly difficult because it requ...

Super excited to share a new preprint from our lab on design of small-molecule binding proteins using neural networks! The paper has a bit of everything. A new graph neural network, new design algorithms, and experimental validation. www.biorxiv.org/content/10.1...
🧵🧪

Guess what? TPS has extended the deadline to March 19 to submit abstracts for poster presentations and speaking opportunities at our 39th Annual Symposium. Join us in San Francisco June 26 - 29 for 3.5 days of scientific talks.
hashtag#proteinscience hashtag#annualsymposium
lnkd.in/g7VKqX7C

Distal mutations enhance catalysis in designed enzymes by facilitating substrate binding and product release The role of amino-acid residues distant from an enzyme's active site in facilitating the complete catalytic cycle—including substrate binding, chemical transformation, and product release—remains poor...

The take-home message? Distal residues actively shape enzyme catalysis. Optimizing them can remove bottlenecks in substrate binding & product release—boosting activity. Want to dive deeper? Read our full study here: www.biorxiv.org/content/10.1...
(6/6)

Molecular dynamics simulations showed that distal mutations enhance active-site accessibility—either by loosening loops covering the active site or widening bottlenecks for substrate entry & product exit. The enzyme breathes more efficiently! 🌬️ (5/6)

Kinetic solvent viscosity effects & stopped-flow experiments showed that distal mutations don’t just tweak structure—they accelerate substrate binding & product release. (4/6)

Crystal structures showed that active-site mutations pre-organize the catalytic machinery. But distal mutations? They subtly tune conformational dynamics—enhancing productive substates & reshaping the energy landscape of the catalytic cycle. (3/6)

We engineered "Core" and "Shell" variants of three evolved Kemp eliminases to dissect the effects of active-site vs. distal mutations. Core mutations dramatically boosted catalysis. Shell mutations alone? Not much—until they worked together in evolved enzymes. 🔍 (2/6)

How do mutations far from an enzyme's active site influence catalysis? 🤔

Part 2: In collaboration with @fraserlab.bsky.social and @silviaosuna.bsky.social, we investigated this question using de novo Kemp eliminases, revealing effects of distal mutations on the catalytic cycle. 🧵 (1/6)

Thank you for the feedback!

Figure from https://www.biorxiv.org/content/10.1101/2025.02.21.639315v1.full.pdf showing how mutations to the active site (labeled "core" relative to the parental, which is labeled "designed"), to second-shell residues (labeled "shell"), or both (labeled "evolved") affect the conformational dynamics of a de novo designed kemp eliminase.

Whereas beneficial active site mutations to enzymes often improve the chemical transformation itself by preorganizing the active site, mutations to second-shell residues instead tune steps like product release by modifying the broader conformational ensemble www.biorxiv.org/content/10.1...

Distal mutations in a designed retro-aldolase alter loop dynamics to shift and accelerate the rate-limiting step Amino-acid residues distant from an enzyme’s active site are known to influence catalysis, but their mechanistic contributions to the catalytic cycle remain poorly understood. Here, we investigate the...

Together, our findings reveal how distal mutations sculpt enzyme function by reshaping the catalytic cycle for more efficient catalysis.

What does this mean for enzyme design? Find out in our preprint: www.biorxiv.org/content/10.1...
#Enzymology #Biophysics #Catalysis #EnzymeDesign

Mechanistically, distal mutations:

🔹 Alter loop flexibility
🔹 Reshape enzyme motions to favor product release
🔹 Realign local electric fields to lower the barrier for C-C bond cleavage

These changes shift the rate-limiting step from C-C bond cleavage to product release—and speed it up!

Crystal structures revealed that distal mutations trigger large-scale conformational shifts in an active-site loop, making the active site more open. Interestingly, these mutations aren't located on the loop—making their effects hard to predict!