Today, we put 2 new #CDlab papers on the @arxiv preprint server – which both report, in different ways, on demonstrating nanoscale rotary motors that are driven by a flow through a nanopore.
@arxiv Such rotors are inspired by the awesome F0F1 ATPase motor protein in our cells. Here, a proton gradient drives rotation of F0 which induces conformational changes in F1 that catalyze production of ATP, which is the fuel for most processes in our body.
Video credit Biovisions
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We built similar rotary motors synthetically from the bottom up, using ‘DNA origami’ in great collaboration with @hendrik_dietz lab. These motor structures dock onto a nanopore and autonomously show sustained unidirectional rotations where a rod rotates at >10 rotations/sec.
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@hendrik_dietz In a first approach (paper 1 arxiv.org/abs/2206.06613), we demonstrate a self-organized artificial nanoscale rotary machine, where a simple DNA bundle acts as the rotor that self organizes onto a nanopore in a thin solid-state membrane that serves as stator and flow generator.
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Video’s often tell the story in less words – so here’s a movie where you see the fluorescently labelled end of a rotating DNA bundle that is observed to move in a circle around the nanopore that is located at the center.
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Energy is provided by applying a static applied DC voltage.
Or even simpler: by merely having different salt concentration on the two sides of the membrane (think salt water versus fresh water)!
The latter directly mimics F0F1 where a concentration gradient drives rotation.
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So how is this rotation driven?
As the elastic and highly charged bundle docks on the pore, it gets compressed by the field into a chiral shape. This asymmetrically couples to the radial water flow out of the pore, which exerts an angular force that drives the rotation.
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Like in this simulation movie…
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We deduced this self-organized rotor mechanism in a wonderful collaboration with the @RGolestanian lab who did extensive simulations on this – see the results for different initial docking configurations where bundles are placed off center in various ways
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@RGolestanian Read all about these self-organized DNA rotors in our preprint (soon to be published in Nature Physics):
In a second paper (arxiv.org/pdf/2206.06612…), advanced rational origami design by the @hendrik_dietz lab defined a true nanoscale turbine, with right-handed or left-handed chiral blades.
Its mere ~20 nm scale mimics the size of the F0F1 ATPase.
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@hendrik_dietz Upon applying a (reinforced 16hb) DNA rod as load, we again measured sustained unidirectional rotations.
And now, the rotation direction was set by chirality! Lefthanded turbines rotated clockwise; righthanded ones rotated anticlockwise
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And, very surprising:
In high salt buffer, the DNA nanoturbines rotated in the opposite direction compared to rotation in low salt!
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Here, full-atom MD simulations by @aksimentievLab (with 4.3 million atoms!) came to the rescue, as they showed the flow-driven rotation as well as the rotation-direction reversal at high salt, caused by a different ion distribution near the DNA that reverses the force on it.
This was all really a technical tour-de-force: from the origami design, to docking turbines correctly, to nanoscale detection, and to puzzling out the mechanisms involved.
Very gratifying that 7 years after the conception of these ideas, this now materialized in 2 papers!
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Our nanoturbines are the first-ever experimental realization of flow-driven rotors at the nanoscale, constituting a new class of molecular motors. These synthetic machines convert energy from a static electrochemical gradient into useful mechanical work.
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These artificial turbines operate autonomously in physiological conditions. Next to better understanding motor proteins such as F0F1 ATPase, the results open new perspectives for engineering active robotics at the nanoscale.
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NB All this was teamwork (see above), but most credits go to 1st author @xinshi_d, who led this challenge as postdoc in our #CDlab
Xin is on the job market now and will continue this work as tenure track assistant professor. I greatly recommend him, so get him while you can! 19/
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I’m thrilled to be able to announce that we were granted a NWO Summit grant entitled ‘Evolving Life from Nonlife’ (EVOLF) with 40 million euro for a 10-year project aimed to cross the gap between non-life and life by assembling a living synthetic cell from lifeless biomolecules!
This project aims to address some of the biggest questions: What is life? How do living systems differ from non-living ones? Can we create living cells from lifeless molecules? EVOLF will take an experimental approach and build synthetic cells from the bottom up, from molecules.
While we already created cellular modules for a minimal genome, metabolism, and cell division, EVOLF will employ AI and directed evolution of synthetic cells to integrate cellular functions into one unified synthetic cell that can autonomously replicate, communicate, and evolve.
DNA loop extrusion by SMC motor proteins is an intriguing process that is the key organizer of our chromosomes
As I explained to you before (see our NAR 2022 paper), magnetic tweezers are a powerful single-molecule method to measure single loop extrusion steps (~40nm, 200bp)
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But here's a new twist!
Today, we put a new #CDlab paper on @biorxivpreprint where we developed a new tweezer assay to directly measure how much supercoiling twist is induced into the extruded loop by an individual SMC in each loop-extrusion step!
@biorxivpreprint We show that all 3 SMC complexes induce the same large negative twist (ie, a linking number change Δ𝐿k of -0.6 at each loop-extrusion step) into the extruded loop, independent of step size.
ATP mutants show that ATP binding is the twist-inducing event during the ATPase cycle 2/
Latest #CDlab paper online now in @ScienceMagazine:
In this important hypothesis paper we sketch a possible mechanistic model for SMC motor proteins that make loops in DNA.
This is a collaborative effort of @HaeringLab, JM-Peters, B-Rowland,. and us
Background of this paper is interesting as the 4 authors met at a Titisee conference. Given that each of us had a favorite model (scrunching, swing&clamp, hold&feed), we disagreed.
But we discussed, ending up with this joint paper on a ‘reel-and-seal’ model that fits most data 2/
Here’s how it works:
SMC binds DNA nontopologically. Upon ATP binding, DNA gets clamped onto the heads, which inserts new DNA into the SMC lumen and stretches the SMC arms. Upon ATP hydrolysis, the newly captured DNA then transfers into the extruded DNA loop, enlarging it.
Yesterday we posted a #CDlab paper on @biorxivpreprint that describes careful single-molecule experiments that measure the supercoiling generated by a RNA polymerase during transcription.
Great work by @JanissenRichard @RomanBarth2 @MincoPolinder JacovdTorre
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@biorxivpreprint @JanissenRichard @RomanBarth2 @MincoPolinder Our work tests the classic twin supercoiling domain model (phrased >50 years ago by Maalφe & Kjeldgaard) that states that positive DNA supercoiling is generated ahead of the RNAP and negative supercoiling in its wake.
The data largely confirm it, but leave 1 huge puzzle!
Protein sequencing is a big deal and goes way beyond DNA sequencing. While we have ~20000 protein-coding genes, we have _millions_ of protein variants, mainly because of post-translational modifications that attach a side group to amino acids.
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Phosphorylation is the most frequent PTM, and of particular interest, as dysregulation of phosphorylation pathways is linked to many diseases including cancers, Parkinson’s, Alzheimer’s, and heart disease.
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We lack techniques to measure such PTMs on the single-molecule level! 😳
=> We need new single-molecule techniques
(Mass spectrometry requires typically more than a billion copies and often struggles to identify the correct position of a PTM between multiple candidate sites)
Today we publish a paper in @ScienceMagazine that expands nanopore readings to the proteome:
a nanopore-based scanner to read off PROTEINS at the single-molecule level! 🤩
Awesome experiments by postdoc Henry Brinkerhoff of our #CDlab, with MD simulations of @aksimentievLab
@ScienceMagazine@aksimentievLab Principle reminds of nanopore DNA sequencing: we draw a peptide through a nanopore with a helicase walking on a lead DNA strand, and then read off ion current step signals as amino acids are blocking the pore.