Designed Arginine-Rich RNA-Binding Peptides with Picomolar Affinity

Arginine-rich peptide motifs (ARMs) capable of binding unique RNA structures play critical roles in transcription, translation, RNA trafficking, and RNA packaging. Bacteriophage ARMs necessary for transcription antitermination bind to distinct boxB RNA hairpin sequences with a characteristic induced α-helical structure. Characterization of ARMs from lambdoid phages reveals that the dissociation constant of the P22 bacteriophage model−antitermination complex (P22_(N21)−P22boxB) is 200 ± 56 pM in free solution at physiologic concentrations of monovalent cation, significantly stronger than previously determined by gel mobility shift and polyacrylamide gel coelectophoresis, and 2 orders of magnitude stronger than the tightest known native ARM−RNA interaction at physiological salt. Here, we use a reciprocal design approach to enhance the binding affinity of two separate α-helical ARM−RNA interactions; one derived from the native λ phage antitermination complex and a second isolated using mRNA display selection experiments targeting boxB RNA

Context and conformation dictate function of a transcription antitermination switch

In bacteriophage λ, transcription elongation is regulated by the N protein, which binds a nascent mRNA hairpin (termed boxB) and enables RNA polymerase to read through distal terminators. We have examined the structure, energetics and in vivo function of a number of N−boxB complexes derived from in vitro protein selection. Trp18 fully stacks on the RNA loop in the wild-type structure, and can become partially or completely unstacked when the sequence context is changed three or four residues away, resulting in a recognition interface in which the best binding residues depend on the sequence context. Notably, in vivo antitermination activity correlates with the presence of a stacked aromatic residue at position 18, but not with N−boxB binding affinity. Our work demonstrates that RNA polymerase responds to subtle conformational changes in cis-acting regulatory complexes and that approximation of components is not sufficient to generate a fully functional transcription switch

Generalized Hartmann-Shack array of dielectric metalens sub-arrays for polarimetric beam profiling

To define and characterize optical systems, obtaining information on the amplitude, phase, and polarization profile of optical beams is of utmost importance. Polarimetry using bulk optics is well established to characterize the polarization state. Recently, metasurfaces and metalenses have successfully been introduced as compact optical components. Here, we take the metasurface concept to the system level by realizing arrays of metalens 2*3 sub-arrays, allowing to determine the polarization profile of an optical beam. We use silicon-based metalenses with a numerical aperture of 0.32 and a mean measured diffraction efficiency in transmission mode of 28% at 1550 nm wavelength. Together with a standard camera recording the array foci, our system is extremely compact and allows for real-time beam diagnostics by inspecting the foci amplitudes. By further analyzing the foci displacements in the spirit of a Hartmann-Shack wavefront sensor, we can simultaneously detect phase-gradient profiles. As application examples, we diagnose the polarization profiles of a radially polarized beam, an azimuthally polarized beam, and of a vortex beam.Comment: 20 pages, 6 figures

Differential Modes of Recognition in N Peptide−BoxB Complexes

N proteins from bacteriophages λ, P22, and φ21 modulate transcription elongation by binding nascent “boxB” mRNA hairpins. This RNA recognition is mediated by N-terminal arginine-rich peptide sequences capable of interacting with their cognate boxB RNA targets. Here, we have analyzed the affinity and specificity of the peptide−RNA interactions that modulate this transcriptional switch. To do this, we constructed a series of peptides based on the wild-type λ, P22, and φ21 N protein binding domains ranging from 11 to 22 residues and analyzed their interactions with the leftward and rightward boxB RNA hairpin targets for all three phage. Binding constant (Kd) values were determined using RNA hairpins labeled with 2-aminopurine (2AP) and monitoring the fluorescence change as peptide was added. Kd's demonstrate that λ and P22 N peptides bind to their cognate boxB targets with high specificity and show equal affinities for their leftward and rightward hairpins. Surprisingly, φ21 shows very little specificity for its cognate targets. λ and P22 N peptides exhibit differential modes of recognition with specificity conferred by their amino- and carboxy-terminal modules, respectively. We have generated a reciprocal matrix of substituted peptides to examine the contributions of individual residues to specificity. Amino acid coupling analysis supports a binding model where the Arg8 residue of λ peptide acts as a conformational hot spot, anchoring the induced loop fold of its boxB hairpin target

Two types of zero Hall phenomena in few-layer MnBi2_2Te4_4

The van der Waals antiferromagnetic topological insulator MnBi$_2$Te$_4$ represents a promising platform for exploring the layer-dependent magnetism and topological states of matter. Despite the realization of several quantized phenomena, such as the quantum anomalous Hall effect and the axion insulator state, the recently observed discrepancies between magnetic and transport properties have aroused controversies concerning the topological nature of MnBi$_2$Te$_4$ in the ground state. Here, we demonstrate the existence of two distinct types of zero Hall phenomena in few-layer MnBi$_2$Te$_4$. In addition to the robust zero Hall plateau associated with the axion insulator state, an unexpected zero Hall phenomenon also occurs in some odd-number-septuple layer devices. Importantly, a statistical survey of the optical contrast in more than 200 MnBi$_2$Te$_4$ reveals that such accidental zero Hall phenomenon arises from the reduction of effective thickness during fabrication process, a factor that was rarely noticed in previous studies of 2D materials. Our finding not only resolves the controversies on the relation between magnetism and anomalous Hall effect in MnBi$_2$Te$_4$, but also highlights the critical issues concerning the fabrication and characterization of devices based on 2D materials.Comment: 21 pages, 4 figure