Polypeptide analysis for nanopore-based protein identification

Presently,proteins are identified by cleaving them with proteases,measuring the mass to charge ratio of the fragments with a mass spectrometer,and matching the fragments to segments within known proteins in databases.We earlier demonstrated that a nanometer-scale pore formed by aerolysin(AeL)can discriminate between,and therefore identify,three similar size proteins from their trypsin-cleaved polypeptide fragments.With this nanopore-protease method,the protein’s identity is instead determined from characteristic ionic current blockade patterns caused by the polypeptide fragments that enter the nanopore.The results also suggested that not all of the theoretically expected cleavage products partition into the pore.To better understand the mechanism by which polypeptide fragments are captured,and how different polypeptides reduce the pore’s ionic current,we studied the effects of 11 identical length polypeptides with different net charges and charge distributions.We show that under certain experimental conditions,negative,positive,and neutral polypeptides are driven into the AeL pore by the same applied voltage polarity.The capture rate and dwell time of polypeptides in the pore depend strongly on the ionic strength,the magnitude of the applied voltage,and the net charge and charge distribution of the polypeptides.The dwell time distribution depends nonmonotonically on the applied voltage(regardless of the polymer’s net charge),and its maximum value depends on the polypeptide net charge and charge distribution.The maximum dwell time for different polypeptides does not occur at the same applied voltage amplitude,which conceivably might complicate the detection and discrimination of some polypeptide fragments.Although additional experiments,computer simulations,and artificial intelligence research are needed to better understand how to optimize the partitioning of enzymatically cleaved fragments into the AeL nanopore,the method is still capable of accurately identifying proteins.

A New Boundary Condition for Three-Dimensional Lattice Boltzmann Simulations of Capillary Filling in Rough Micro-Channels

A new boundary condition,aimed at inhibiting near-wall condensation effects in lattice Boltzmann simulations of capillary flows in micro-corrugated channels,is introduced.The new boundary condition is validated against analytical solutions for smooth channels and demonstrated for the case of three-dimensional microflows over randomly corrugated walls.

Effacements of Form

The theme of this essay is expressed in a line from the Dao De Jing： ＂The great image has no form.＂ The essay shows how this effacement, annulment, or withdrawal of form is realized in ancient Chinese painting （Song and Ming Dynasties） and in the conception of the natural elements to which much of this painting is related. Certain resonances with this effacement of form are identified in the way that recent Continental thought focuses on an effacement of form as it was determined in ancient Greek philosophy.

Prospective Merger Between Car-Parrinello and Lattice Boltzmann Methods for Quantum Many-Body Simulations

Formal analogies between the Car-Parrinello(CP)ab-initio molecular dynamics for quantum many-body systems,and the Lattice Boltzmann(LB)method for classical and quantum fluids,are pointed out.A theoretical scenario,whereby the quantum LB would be coupled to the CP framework to speed-up many-body quantum simulations,is also discussed,together with accompanying considerations on the computational efficiency of the prospective CP-LB scheme.

Red Light-Induced Phytochrome Relocation into the Nucleus in Adiantum capillus-veneris

Phytochromes in seed plants are known to move into nuclei in a red light-dependent manner with or without interacting factors. Here, we show phytochrome relocation to the nuclear region in phytochrome-dependent Adiantum capillus-veneris spore germination by partial spore-irradiation experiments. The nuclear or non-nuclear region of imbibed spores was irradiated with a microbeam of red and/or far-red light and the localization of phytochrome involved in spore germination was estimated from the germination rate. The phytochrome for spore germination existed throughout whole spore under darkness after imbibition, but gradually migrated to the nuclear region following red light irradiation. In- tracellular distribution of PHY-GUS fusion proteins expressed in germinated spores by particle bombardment showed the migration of Acphy2, but not Acphyl, into nucleus in a red light-dependent manner, suggesting that Acphy2 is the photoreceptor for fern spore germination.

DICER-LIKE3 Activity in Physcomitrella patens DICER-LIKE4 Mutants Causes Severe Developmental Dysfunction and Sterility

Trans-acting small interfering RNAs （ta-siRNAs） are plant-specific siRNAs released from TAS precursor tran- scripts after microRNA-dependent cleavage, conversion into double-stranded RNA, and Dicer-dependent phased process- ing. Like microRNAs （miRNAs）, ta-siRNAs direct site-specific cleavage of target RNAs at sites of extensive complementarity. Here, we show that the DICER-LIKE 4 protein of Physcomitrella patens （PpDCL4） is essential for the biogenesis of 21 nucleotide （nt） ta-siRNAs. In APpDCL4 mutants, off-sized 23 and 24-nt ta-siRNAs accumulated as the result of PpDCL3 activity. APpDCL4 mutants display severe abnormalities throughout Physcomitrella development, including sterility, that were fully reversed in APpDCL3/APpDCL4 double-mutant plants. Therefore, PpDCL3 activity, not loss of PpDCL4 function per se, is the cause of the APpDCL4 phenotypes. Additionally, we describe several new Physcomitrella trans-acting siRNA loci, three of which belong to a new family, TA56. TAS6 loci are typified by sliced miR156 and miR529 target sites and are in close proximity to PpTAS3 loci.

Conserved Cytochrome P450 Evolved in Seed Plants Regulates Flower Maturation

Global inspection of plant genomes identifies genes maintained in low copies across taxa and under strong purifying selection, which are likely to have essential functions. Based on this rationale, we investigated the function of the low-duplicated CYP715 cytochrome P450 gene family that appeared early in seed plants and evolved under strong negative selection. Arabidopsis CYP715A 1 showed a restricted tissue-specific expres- sion in the tapetum of flower buds and in the anther filaments upon anthesis, cyp715a1 insertion lines showed a strong defect in petal development, and transient alteration of pollen intine deposition. Comparative expres- sion analysis revealed the downregulated expression of genes involved in pollen development, cell wall biogenesis, hormone homeostasis, and floral sesquiterpene biosynthesis, especially TPS21 and several key genes regulating floral development such as MYB21, MYB24, and MYC2. Accordingly, floral sesquiterpene emission was suppressed in the cyp715a1 mutants. Flower hormone profiling, in addition, indicated a modi- fication of gibberellin homeostasis and a strong disturbance of the turnover of jasmonic acid derivatives. Petal growth was partially restored by the active gibberellin GA3 or the functional analog of jasmonoyl-isoleucine, coronatine. CYP715 appears to function as a key regulator of flower maturation, synchronizing petal expan- sion and volatile emission. It is thus expected to be an important determinant of flower-insect interaction.

The role of weather and density dependence on population dynamics of Alpine-dwelling red deer

The dynamics of red deer Cervus elaphus populations has been investigated across different environmental conditions,with the notable exception of the European Alps.Although the population dynamics of mountain-dwelling ungulates is typically influenced by the interaction between winter severity and density,the increase of temperatures and the reduction of snowpack occurring on the Alps since the 1980s may be expected to alter this pattern,especially in populations dwelling at medium-low elevations.Taking advantage of a 29-year time series of spring count data,we explored the role of weather stochasticity and density dependence on growth rate and vital rates(mortality and weaning success),and the density-dependent variation in body mass in a red deer population of the Italian Alps.The interaction between increasing values of density and snow depth exerted negative and positive effects on growth and mortality rates,respectively,while weaning success was negatively affected by increasing values of density,female-biased sex ratio and snow depth.Body mass of males and females of different age classes declined as population size increased.Our data support the role of winter severity and density dependence as key components of red deer population dynamics,and provide insight into the species’ecology on the European Alps.Despite the recent decline of snowpack on the Alpine Region,the negative impacts of winter severity and population abundance on growth rrate(possibly mediated by the density-dependent decline in body mass)confirms the importance of overwinter mortality in affecting the population dynamics of Alpine-dwelling red deer.

Multiscale Hemodynamics Using GPU Clusters

The parallel implementation of MUPHY,a concurrent multiscale code for large-scale hemodynamic simulations in anatomically realistic geometries,for multi-GPU platforms is presented.Performance tests show excellent results,with a nearly linear parallel speed-up on up to 32GPUs and a more than tenfold GPU/CPU acceleration,all across the range of GPUs.The basic MUPHY scheme combines a hydrokinetic(Lattice Boltzmann)representation of the blood plasma,with a Particle Dynamics treatment of suspended biological bodies,such as red blood cells.To the best of our knowledge,this represents the first effort in the direction of laying down general design principles for multiscale/physics parallel Particle Dynamics applications in non-ideal geometries.This configures the present multi-GPU version of MUPHY as one of the first examples of a high-performance parallel code for multiscale/physics biofluidic applications in realistically complex geometries.

Molecular Dynamics Simulations of Nanoparticle Interactions with a Planar Wall:Does Shape Matter?

We investigate the hydrodynamic interactions of spherical colloidal nano particles and nano tetrahedra near a planar wall by means of molecular dynamics(MD)simulations of rigid particles within an all-atom solvent.For both spherical and nano-tetrahedral particles,we find that the parallel and perpendicular components of the local diffusion coefficient and viscosity,show good agreement with hydrodynamic theory of Fax´en and Brenner.This provides further evidence that low perturbations from sphericality of a nanoparticle’s shape has little influence on its local diffusive behaviour,and that for this particular case,the continuum theory fluid dynamics is valid even down to molecular scales.