My invited review “Peptoids at the 7th Summit: Toward Macromolecular Systems Engineering” [pdf] kicks off the peptoid special issue of Biopolymers: Peptide Science.

Astoundingly, all the papers are open access.

Here’s the abstract:

Peptoids at the 7th Summit:
Toward Macromolecular Systems Engineering

Methods for facile synthesis of extraordinarily diverse peptide-like oligomers have placed peptoids at the center of a broad and vibrant area of foldamer science and technology. The 7th Peptoid Summit offered a perspective on the current state of peptoid science and technology and on prospects for engineering supramolecular assemblies that rival the complexity of biomolecular systems. Methods for engineering biomolecular systems based on DNA and protein are advancing rapidly, building a technology platform for engineering increasingly large and complex self-assembled nanosystems. A comparative review of the physical basis for DNA, protein, and peptoid engineering indicates that the characteristics of peptoids suit them for a strong role in developing self-assembled nanosystems. Physical parallels between peptoids and proteins indicate that peptoid engineering, like protein engineering, will require specialized software to support design. Access to novel side-chain functionality will enable peptoid designers to exploit novel binding interactions, including many that have been discovered and exploited in crystal engineering, a field that has extensively explored the self-assembly of small organic molecules to form well-ordered structures. Developments in DNA, protein, and inorganic nanotechnologies are converging to provide a technology platform for the design and fabrication of complex, functional, atomically precise nanosystems. Peptoid-based foldamer technologies can contribute to this convergence, expanding the scope of the emerging field of atomically precise macromolecular nanosystems.

As you can see, the paper goes beyond peptoids (a favorite topic here) to examine the broader context of foldamer design for engineering atomically precise systems. Read it for a summary of some important developments and prospects.

A paper in Science reports a design method that substantially advances the macromolecular technology base for building atomically precise nanosystems.

Background: foldamer engineering

As many readers know, biology shows an effective way build large, intricate, atomically precise systems: Use covalent chemistry to build chains of small building blocks, and design these chains to fold into nanoscale building blocks that undergo spontaneous assembly driven by Brownian motion and selective binding. This is a key step in climbing a ladder of fabrication technologies that leads to broader, more powerful capabilities.

The covalent synthesis of suitable chains of building blocks* was mastered decades ago, using programmable nanoscale machines that operate in biological systems. Designing structures that fold into compact nanoscale objects has become increasingly routine. Designing these building blocks to assemble, however, has lagged.

The approach

This highlights the importance of the paper in Science.

The authors (from the Baker lab, and I’m tempted to add “of course”) used RosettaDesign-based protein engineering tools to design proteins with surface structures that bind to a natural protein at a particular location, and with a particular orientation. Finding a protein that binds isn’t too hard — screening and evolutionary methods applied to antibodies (among other proteins) can do this — but achieving high affinity (tight binding) in a specific geometry is new.

They achieved this by designing binders with the correct geometry but mediocre binding, and then using selection (the equivalent of antibody affinity maturation) to refine the interfaces to achieve high affinity. The refinement process retains the initial alignment with good fidelity.

The binding target was a conserved region of the influenza hemagglutinin molecule, hinting at an approach to developing a subtype-independent anti-influenza therapy.

Solving a harder problem than necessary

Note, however, that authors didn’t address the problem of designing building-block interfaces, as an engineer would understand the task: They did something harder. Only side of the interface was designed to bind, while the other was a naturally occurring structure that normally binds nothing.
An engineer designing building-block assemblies, by contrast, would design the interface as a unit, not just one side of it.

It’s easy to see the advantages of being free to tweak both sides to achieve a good fit, to balance solubility and costs of desolvation, and to introduce specific binding interactions (hydrogen bonds, salt bridges, hydrophobic pockets on one side that match hydrophobic side chains on the other, etc.). Freedom to design both sides together also means that protein engineers — when pursuing engineering objectives — can exploit the best-understood motifs, rather than deliberately plunging into the unknown.

In conventional engineering, no one designing a system would freeze the design of one component, and then attempt to mate another to it at a location not designed for the purpose. Interfaces aren’t afterthoughts.

A companion perspective piece for the paper observes that

Although Fleishman et al. have produced a landmark result, it is evident that computational protein interface design is not a solved problem.

For the more symmetric engineering design problem, however, the methods described in the paper can be expected to provide a basis for reliable design tools.

I look forward to seeing the methods and the lab results. This should be low-hanging fruit.

* In other words, peptide foldamers (commonly called “proteins”) which include a range of high-performance engineering polymers.

See also:

• A High-Performance Polymer for Nanosytems Engineering
• Macromolecular Modeling for Molecular Systems Engineering
• The Molecular Machine Path to Molecular Manufacturing (1):
  Foldamers and Brownian Assembly
• The Antiparallel Structures of Science and Engineering

Background: polyoxometalate nanostructures are cool (more here).

Lee Cronin sent me a pdf of the polyoxometalate paper I discussed in my previous post, and he notes that readers can download it here, with other papers on his group’s website here.

The Israel Journal of Chemistry has a new special issue, “Frontiers in Metal Oxide Cluster Science”, that includes an overview with the provocative title “Oxo-Metalate Building Blocks: Conceptual Competitors for Tetravalent Carbon?” [pdf]. Excerpt:

These developments offer options for controlling structure and function—in principle analogous to activities in organic chemistry.

Note that part of the story is bridging POM chemistry to organic chemistry itself.

In this connection, chemists may find this Chemical Communications review particularly interesting: “Functionalization of polyoxometalates: towards advanced applications in catalysis and materials science”. I’d intended to post a link to the pdf, but the free version has apparently disappeared.

(By the way, in exploring the literature across a range of disciplines, I’ve found that chemistry journals are among the worst for open access. Even my ACS membership gives me online access to exactly zero ACS journals — and members are starting to complain.)

New work with exosomes promises wide-ranging advances in medicine, courtesy of an emerging biomolecular nanotechnology.

As pharmaceutical chemists know, the blood-brain barrier blocks delivery of many molecules that do wonderful things if injected directly into the brain, but injecting the brain isn’t quite as convenient as injecting a vein.

Exosomes are lipid vesicles manufactured by cells for transporting diverse molecules to other cells, including signaling molecules such as micro RNAs. Now, they’ve been shown to carry their contents across the blood-brain barrier, and other work has shown that exosome-like particles can be made synthetically, with membranes chock-full of functional molecules for targeting cells and inducing responses from them. With diameters of 30 to 100 nm, exosomes have room for a lot of payload.

BBC report here: “Breakthrough in delivering drugs to the brain”, abstract of paper in Nature Biotechnology here: “Delivery of siRNA to the mouse brain by systemic injection of targeted exosomes”.

The biomedical potential of siRNA is enormous, and delivery has been the main obstacle to wide-ranging applications.

Image and atomic reconstruction

From the abstract:

Although atomic-resolution electron microscopy has been feasible for nearly four decades, neither electron tomography nor any other experimental technique has yet demonstrated atomic resolution in three dimensions. Here we report the 3D reconstruction of a complex crystalline nanoparticle at atomic resolution. To achieve this, we combined aberration-corrected scanning transmission electron microscopy, statistical parameter estimation theory, and discrete tomography, Unlike conventional electron tomography, only two images of the target—a silver nanoparticle embedded in an aluminium matrix—are sufficient for the reconstruction when combined with available knowledge about the particle’s crystallographic structure. Additional projections confirm the reliability of the result. The results we present help close the gap between the atomic resolution achievable in two-dimensional electron micrographs and the coarser resolution that has hitherto been obtained by conventional electron tomography. [Links added]

This computational reconstruction technique (imaging aided by inference) requires only two images, but it relies on prior knowledge (or assumptions) about the structure: for example, that the crystal is face-centered cubic structure, has no holes, and no deep grooves in its surface. The data analysis uses a stochastic method (simulated annealing), and 16 independent reconstruction runs gave a difference in the positioning of 41 (out of 784) atoms in the specimen, so the result isn’t fully determined.

When a structure results from what should be an atomically precise process, knowledge (or reasonable expectations) will typically be extensive and detailed, leaving only narrow questions to be answered by imaging — the orientation of parts across an interface, for example, or the nature of a defect in a mostly-correct structure. Imaging aided by inference should again be quite powerful, and helpful in debugging fabrication processes.

The specimen in the current study is a silver nanoparticle in an aluminum matrix, that is, an array of electron-dense atoms in a radiation-tolerant structure. Studies of biomolecular structures still face the problems of radiation damage to delicate structures that provide lower contrasts in electron density, but cryogenic electron tomography has been advancing in this domain, too. Here, covalent structures with local conformational constraints are typically known at the outset, and this has been used to provide a bridge from low-resolution images to inferred (almost) atomically precise structures.

• Electron cryomicroscopy reaches landmark molecular resolution

This news just in:

A ‘buckyball’ — a spherical molecule made up of 60 carbon atoms — has been turned into a vial just big enough to hold a single water molecule….

The authors say that uses for the vial could include acting as a carrier for drugs in the body.
(News item)

As The Onion might ask, What do you think?

As I’ve said, scientists are held to a high standard when talking about scientific results with their peers, and a noticeably different standard when talking about potential applications with everyone else.

This just might be a cumulative problem for the credibility of science regarding (for example) climate change, and maybe even the potential of nanotechnology.

Compared to small molecules, nanoparticles offer more physical scope for functional engineering, and according to a report in Science, more than 50 companies are pressing forward to exploit this for cancer diagnosis and treatment. Nearly a dozen nanoparticle-based medicines are reportedly in clinical trials, and lab research suggests a road to programmable control of cellular functions.

Nanoparticles make good delivery vehicles for molecular cargo in vivo. The have distinct surfaces and interiors, and this makes it possible separately engineer for solubility, immunological compatibility, targeting and penetration of cells, and controlled release of compounds that, when simply injected, are ineffective or horribly toxic.

Not just size and surface properties matter: Shape makes a difference (cylinders penetrate cells better than spheres), and even mechanical stiffness is important — soft particles can remain in circulation for almost 100 hours, where rigid particles are cleared in 1/50 the time.

Looking forward

The advantages of nanoscale engineering for biomedicine will increase in concert with increasing capabilities in molecular and supramolecular engineering. There’s a continuum of prospective technologies that stretches from macromolecular conjugation and liposome encapsulation of drugs, through applications of current-generation multifunctional liposomes and nanoparticles, to medical interventions based on particles that undergo differential activation in response to weighted molecular sensing inputs and threshold measurements, evaluated by Boolean logic*. And then more, and more beyond that.

(* Programmable intracellular Boolean logic has already been demonstrated: see below.)

The wonders of siRNA and the role of nanoparticles

Keep a keen eye on advances in small interfering RNA (siRNA) technology: By modulating protein expression, siRNA therapeutics promise to provide remarkably flexible and specific control of cellular processes. RNA molecules have have short half-lives in circulation and don’t readily enter cells, but engineered nanoparticles promise to overcome this obstacle. (Recent research abstract & pdf; recent review abstract & pdf.)

Biocompatible molecular Boolean logic has already been demonstrated, with RNA as the basis:

“A universal RNAi-based logic evaluator
that operates in mammalian cells”

Abstract, Nature Biotechnology:
…The encoding rules, combined with a specific arrangement of the siRNA targets in a synthetic gene network, allow direct evaluation of any Boolean expression in standard forms using siRNAs and indirect evaluation using endogenous inputs. We demonstrate direct evaluation of expressions with up to five logic variables. Implementation of the encoding rules through sensory up- and down-regulatory links between the inputs and siRNA mediators will allow arbitrary Boolean decision-making using these inputs.

PDF here.

Electron microscopes can image biological macromolecules in cryogenic ice, but it shows them as low-contrast features in a grainy image (see below). Using enough electrons to reduce the graininess would first destroy the specimen.

The trick to getting enough information without frying the molecules is to image many specimens that are known to be identical, and to somehow find, align, and combine data from their images (keep in mind that these are grainy, 2D shadows of randomly oriented 3D objects). The quality of the resulting reconstruction depends heavily on the quality of the methods, and the methods are not simple.

A new report in Nature describes improved methods and a result (below) that sharpens resolution from the previous 1.3 nm to 0.66 nm. This provides about 8 times as many voxels, and gave the authors enough information to infer protein secondary structures and build a reliable atomic model of F-actin:

Raw electron cryomicroscopy images,
F-actin helical assembly.

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Reconstruction at 0.66 nm resolution.
(Stereo view of density isosurface with fitted C? ribbon diagram.)

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“Direct visualization of secondary structures of F-actin by electron cryomicroscopy”,
Takashi Fujii, et al., Nature 467:724–728 (2010).

The authors’ method is apparently quite practical. From the article:

We have demonstrated that our cryoEM technologies now allow us to visualize the secondary structures of such thin objects as F-actin in a few days of work, including image data collection and processing and 3D image reconstruction.

In closing, they remark that

There is also room for further improvement, to reach atomic resolution. The present work offers a new opportunity to look into cellular mechanisms essential for the activities of life.

And in building nonbiological molecular machinery, of course, it’s useful to be able to see the result.

• A DNA-Imaging Bottleneck
• Graphene Nanotechnology (and TEAM Microscopes)
• Molecular Electron Holography: Progress toward atomic-resolution imaging?
• A Third Revolution in DNA Nanotechnology