A Bipedal DNA Brownian Motor with Coordinated Legs

Science, 2009, 324, 67

A substantial challenge in engineering molecular motors is designing mechanisms to coordinate the motion between multiple domains of the motor so as to bias random thermal motion. For bipedal motors, this challenge takes the form of coordinating the movement of the biped’s legs so that they can move in a synchronized fashion. To address this problem, we have constructed an autonomous DNA bipedal walker that coordinates the action of its two legs by cyclically catalyzing the hybridization of metastable DNA fuel strands. This process leads to a chemically ratcheted walk along a directionally polar DNA track. By covalently cross-linking aliquots of the walker to its track in successive walking states, we demonstrate that this Brownian motor can complete a full walking cycle on a track whose length could be extended for longer walks. We believe that this study helps to uncover principles behind the design of unidirectional devices that can function without intervention. This device should be able to fulfill roles that entail the performance of useful mechanical work on the nanometer scale.

Photodegradable Hydrogels for Dynamic Tuning of Physical and Chemical Properties

Science, 2009, 324, 59

We report a strategy to create photodegradable poly(ethylene glycol)–based hydrogels through rapid polymerization of cytocompatible macromers for remote manipulation of gel properties in situ. Postgelation control of the gel properties was demonstrated to introduce temporal changes, creation of arbitrarily shaped features, and on-demand pendant functionality release. Channels photodegraded within a hydrogel containing encapsulated cells allow cell migration. Temporal variation of the biochemical gel composition was used to influence chondrogenic differentiation of encapsulated stem cells. Photodegradable gels that allow real-time manipulation of material properties or chemistry provide dynamic environments with the scope to answer fundamental questions about material regulation of live cell function and may affect an array of applications from design of drug delivery vehicles to tissue engineering systems.

Structure of P-Glycoprotein Reveals a Molecular Basis for Poly-Specific Drug Binding

Science, 2009, 323, 1718

P-glycoprotein (P-gp) detoxifies cells by exporting hundreds of chemically unrelated toxins but has been implicated in multidrug resistance (MDR) in the treatment of cancers. Substrate promiscuity is a hallmark of P-gp activity, thus a structural description of poly-specific drug-binding is important for the rational design of anticancer drugs and MDR inhibitors. The x-ray structure of apo P-gp at 3.8 angstroms reveals an internal cavity of 6000 angstroms cubed with a 30 angstrom separation of the two nucleotide-binding domains. Two additional P-gp structures with cyclic peptide inhibitors demonstrate distinct drug-binding sites in the internal cavity capable of stereoselectivity that is based on hydrophobic and aromatic interactions. Apo and drug-bound P-gp structures have portals open to the cytoplasm and the inner leaflet of the lipid bilayer for drug entry. The inward-facing conformation represents an initial stage of the transport cycle that is competent for drug binding.

A Role for RNAi in the Selective Correction of DNA Methylation Defects

Science, 2009, 323, 1600

DNA methylation is essential for silencing transposable elements and some genes in higher eukaryotes, which suggests that this modification must be tightly controlled. However, accidental changes in DNA methylation can be transmitted through mitosis (as in cancer) or meiosis, leading to epiallelic variation. We demonstrated the existence of an efficient mechanism that protects against transgenerational loss of DNA methylation in Arabidopsis. Remethylation is specific to the subset of heavily methylated repeats that are targeted by the RNA interference (RNAi) machinery. This process does not spread into flanking regions, is usually progressive over several generations, and faithfully restores wild-type methylation over target sequences in an RNAi-dependent manner. Our findings suggest an important role for RNAi in protecting genomes against long-term epigenetic defects.

Variants of the Antibody Herceptin That Interact with HER2 and VEGF at the Antigen Binding Site

Science, 2009, 323, 1610

The interface between antibody and antigen is often depicted as a lock and key, suggesting that an antibody surface can accommodate only one antigen. Here, we describe an antibody with an antigen binding site that binds two distinct proteins with high affinity. We isolated a variant of Herceptin, a therapeutic monoclonal antibody that binds the human epidermal growth factor receptor 2 (HER2), on the basis of its ability to simultaneously interact with vascular endothelial growth factor (VEGF). Crystallographic and mutagenesis studies revealed that distinct amino acids of this antibody, called bH1, engage HER2 and VEGF energetically, but there is extensive overlap between the antibody surface areas contacting the two antigens. An affinity-improved version of bH1 inhibits both HER2- and VEGF-mediated cell proliferation in vitro and tumor progression in mouse models. Such “two-in-one” antibodies challenge the monoclonal antibody paradigm of one binding site, one antigen. They could also provide new opportunities for antibody-based therapy.

Giant-Stroke, Superelastic Carbon Nanotube Aerogel Muscles

Science, 2009, 323, 1575

Improved electrically powered artificial muscles are needed for generating force, moving objects, and accomplishing work. Carbon nanotube aerogel sheets are the sole component of new artificial muscles that provide giant elongations and elongation rates of 220% and (3.7 x 104)% per second, respectively, at operating temperatures from 80 to 1900 kelvin. These solid-state–fabricated sheets are enthalpic rubbers having gaslike density and specific strength in one direction higher than those of steel plate. Actuation decreases nanotube aerogel density and can be permanently frozen for such device applications as transparent electrodes. Poisson’s ratios reach 15, a factor of 30 higher than for conventional rubbers. These giant Poisson’s ratios explain the observed opposite sign of width and length actuation and result in rare properties: negative linear compressibility and stretch densification.

Variants of the Antibody Herceptin That Interact with HER2 and VEGF at the Antigen Binding Site

Science, 2009, 323, 1610

The interface between antibody and antigen is often depicted as a lock and key, suggesting that an antibody surface can accommodate only one antigen. Here, we describe an antibody with an antigen binding site that binds two distinct proteins with high affinity. We isolated a variant of Herceptin, a therapeutic monoclonal antibody that binds the human epidermal growth factor receptor 2 (HER2), on the basis of its ability to simultaneously interact with vascular endothelial growth factor (VEGF). Crystallographic and mutagenesis studies revealed that distinct amino acids of this antibody, called bH1, engage HER2 and VEGF energetically, but there is extensive overlap between the antibody surface areas contacting the two antigens. An affinity-improved version of bH1 inhibits both HER2- and VEGF-mediated cell proliferation in vitro and tumor progression in mouse models. Such “two-in-one” antibodies challenge the monoclonal antibody paradigm of one binding site, one antigen. They could also provide new opportunities for antibody-based therapy.

The Dynamic Control of Kiss-And-Run and Vesicular Reuse Probed with Single Nanoparticles

Science, 2009, 323, 1448

Vesicular secretion of neurotransmitter is essential for neuronal communication. Kiss-and-run is a mode of membrane fusion and retrieval without the full collapse of the vesicle into the plasma membrane and de novo regeneration. The importance of kiss-and-run during efficient neurotransmission has remained in doubt. We developed an approach for loading individual synaptic vesicles with single quantum dots. Their size and pH-dependent photoluminescence change allowed us to distinguish kiss-and-run from full-collapse fusion and to track single vesicles through multiple rounds of kiss-and-run and reuse, without perturbing vesicle cycling. Kiss-and-run dominated at the beginning of stimulus trains, reflecting the preference of vesicles with high release probability. Its incidence was increased by rapid firing, a response appropriate to shape the kinetics of neurotransmission during a wide range of firing patterns.

Brightly Fluorescent Single-Walled Carbon Nanotubes via an Oxygen-Excluding Surfactant Organization

Science, 2009, 323, 1319

Attaining high photoluminescence quantum yields for single-walled carbon nanotubes (SWNTs) in order to broaden their optoelectronics and sensing applications has been a challenging task. Among various nonradiative pathways, sidewall chemisorption of oxygen provides a known defect for exciton quenching through nanotube hole doping. We found that an aliphatic (dodecyl) analog of flavin mononucleotide, FC12, leads to high dispersion of SWNTs, which tend to aggregate into bundles. Unlike other surfactants, the surface organization of FC12 is sufficiently tight to exclude oxygen from the SWNT surface, which led to quantum yields as high as 20%. Toluene-dispersed, FC12-wrapped nanotubes exhibited an absorption spectrum with ultrasharp peaks (widths of 12 to 25 milli–electron volts) devoid of the characteristic background absorption of most nanotube dispersions.

Self-Sustained Replication of an RNA Enzyme

Science, 2009, 323, 1229

An RNA enzyme that catalyzes the RNA-templated joining of RNA was converted to a format whereby two enzymes catalyze each other’s synthesis from a total of four oligonucleotide substrates. These cross-replicating RNA enzymes undergo self-sustained exponential amplification in the absence of proteins or other biological materials. Amplification occurs with a doubling time of about 1 hour and can be continued indefinitely. Populations of various cross-replicating enzymes were constructed and allowed to compete for a common pool of substrates, during which recombinant replicators arose and grew to dominate the population. These replicating RNA enzymes can serve as an experimental model of a genetic system. Many such model systems could be constructed, allowing different selective outcomes to be related to the underlying properties of the genetic system.