Visualization of a water-selective pore by electron crystallography in vitreous ice

The water-selective pathway through the aquaporin-1 membrane channel has been visualized by fitting an atomic model to a 3.7-Å resolution three-dimensional density map. This map was determined by analyzing images and electron diffraction patterns of lipid-reconstituted two-dimensional crystals of aquaporin-1 preserved in vitrified buffer in the absence of any additive. The aqueous pathway is characterized by a size-selective pore that is ≈4.0 ± 0.5Å in diameter, spans a length of ≈18Å, and bends by ≈25° as it traverses the bilayer. This narrow pore is connected by wide, funnel-shaped openings at the extracellular and cytoplasmic faces. The size-selective pore is outlined mostly by hydrophobic residues, resulting in a relatively inert pathway conducive to diffusion-limited water flow. The apex of the curved pore is close to the locations of the in-plane pseudo-2-fold symmetry axis that relates the N- and C-terminal halves and the conserved, functionally important N76 and N192 residues.

Design of three-dimensional domain-swapped dimers and fibrous oligomers

Three-dimensional (3D) domain-swapped proteins are intermolecularly folded analogs of monomeric proteins; both are stabilized by the identical interactions, but the individual domains interact intramolecularly in monomeric proteins, whereas they form intermolecular interactions in 3D domain-swapped structures. The structures and conditions of formation of several domain-swapped dimers and trimers are known, but the formation of higher order 3D domain-swapped oligomers has been less thoroughly studied. Here we contrast the structural consequences of domain swapping from two designed three-helix bundles: one with an up-down-up topology, and the other with an up-down-down topology. The up-down-up topology gives rise to a domain-swapped dimer whose structure has been determined to 1.5 Å resolution by x-ray crystallography. In contrast, the domain-swapped protein with an up-down-down topology forms fibrils as shown by electron microscopy and dynamic light scattering. This demonstrates that design principles can predict the oligomeric state of 3D domain-swapped molecules, which should aid in the design of domain-swapped proteins and biomaterials.

Three-dimensional image reconstruction of dephosphorylated smooth muscle heavy meromyosin reveals asymmetry in the interaction between myosin heads and placement of subfragment 2

Regulation of the actin-activated ATPase of smooth muscle myosin II is known to involve an interaction between the two heads that is controlled by phosphorylation of the regulatory light chain. However, the three-dimensional structure of this inactivated form has been unknown. We have used a lipid monolayer to obtain two-dimensional crystalline arrays of the unphosphorylated inactive form of smooth muscle heavy meromyosin suitable for structural studies by electron cryomicroscopy of unstained, frozen-hydrated specimens. The three-dimensional structure reveals an asymmetric interaction between the two myosin heads. The ATPase activity of one head is sterically “blocked” because part of its actin-binding interface is positioned onto the converter domain of the second head. ATPase activity of the second head, which can bind actin, appears to be inhibited through stabilization of converter domain movements needed to release phosphate and achieve strong actin binding. When the subfragment 2 domain of heavy meromyosin is oriented as it would be in an actomyosin filament lattice, the position of the heads is very different from that needed to bind actin, suggesting an additional contribution to ATPase inhibition in situ.

Novel phenomena in dilute electron systems in two dimensions

For the past two decades, all two-dimensional systems of electrons were believed to be insulating in the limit of zero temperature. Recent experiments provide evidence for an unexpected transition to a conducting phase at very low electron densities. The nature of this phase is not understood and is currently the focus of intense theoretical and experimental attention.

An approach to three-dimensional structures of biomolecules by using single-molecule diffraction images

We describe an approach to the high-resolution three-dimensional structural determination of macromolecules that utilizes ultrashort, intense x-ray pulses to record diffraction data in combination with direct phase retrieval by the oversampling technique. It is shown that a simulated molecular diffraction pattern at 2.5-Å resolution accumulated from multiple copies of single rubisco biomolecules, each generated by a femtosecond-level x-ray free electron laser pulse, can be successfully phased and transformed into an accurate electron density map comparable to that obtained by more conventional methods. The phase problem is solved by using an iterative algorithm with a random phase set as an initial input. The convergence speed of the algorithm is reasonably fast, typically around a few hundred iterations. This approach and phasing method do not require any ab initio information about the molecule, do not require an extended ordered lattice array, and can tolerate high noise and some missing intensity data at the center of the diffraction pattern. With the prospects of the x-ray free electron lasers, this approach could provide a major new opportunity for the high-resolution three-dimensional structure determination of single biomolecules.

Cryo-electron microscopy studies of empty capsids of human parvovirus B19 complexed with its cellular receptor.

The three-dimensional structures of human parvovirus B19 VP2 capsids, alone and complexed with its cellular receptor, globoside, have been determined to 26 resolution. The B19 capsid structure, reconstructed from cryo-electron micrographs of vitrified specimens, has depressions on the icosahedral 2-fold and 3-fold axes, as well as a canyon-like region around the 5-fold axes. Similar results had previously been found in an 8 angstrom resolution map derived from x-ray diffraction data. Other parvoviral structures have a cylindrical channel along the 5-fold icosahedral axes, whereas density covers the 5-fold axes in B19. The glycolipid receptor molecules bind into the depressions on the 3-fold axes of the B19:globoside complex. A model of the tetrasaccharide component of globoside, organized as a trimeric fiber, fits well into the difference density representing the globoside receptor. Escape mutations to neutralizing antibodies map onto th capsid surface at regions immediately surrounding the globoside attachment sites. The proximity of the antigenic epitopes to the receptor site suggests that neutralization of virus infectivity is caused by preventing attachment of viruses to cells.

Two-dimensional protein crystallization via metal-ion coordination by naturally occurring surface histidines.

A powerful and potentially general approach to the targeting and crystallization of proteins on lipid interfaces through coordination of surface histidine residues to lipid-chelated divalent metal ions is presented. This approach, which should be applicable to the crystallization of a wide range of naturally occurring or engineered proteins, is illustrated here by the crystallization of streptavidin on a monolayer of an iminodiacetate-Cu(II) lipid spread at the air-water interface. This method allows control of the protein orientation at interfaces, which is significant for the facile production of highly ordered protein arrays and for electron density mapping in structural analysis of two-dimensional crystals. Binding of native streptavidin to the iminodiacetate-Cu lipids occurs via His-87, located on the protein surface near the biotin binding pocket. The two-dimensional streptavidin crystals show a previously undescribed microscopic shape that differs from that of crystals formed beneath biotinylated lipids.

Polarized interacting exciton gas in quantum wells and bulk semiconductors

We develop a theory to calculate exciton binding energies of both two- and three-dimensional spin polarized exciton gases within a mean field approach. Our method allows the analysis of recent experiments showing the importance of the polarization and intensity of the excitation light on the exciton luminescence of GaAs quantum wells. We study the breaking of the spin degeneracy observed at high exciton density (5×1010 cm2). Energy level splitting between spin +1 and spin -1 is shown to be due to many-body interexcitonic exchange while the spin relaxation time is controlled by intraexciton exchange. © 1996 The American Physical Society.

Spin degree of freedom in two dimensional exciton condensates

We present a theoretical analysis of a spin-dependent multicomponent condensate in two dimensions. The case of a condensate of resonantly photoexcited excitons having two different spin orientations is studied in detail. The energy and the chemical potentials of this system depend strongly on the spin polarization. When electrons and holes are located in two different planes, the condensate can be either totally spin polarized or spin unpolarized, a property that is measurable. The phase diagram in terms of the total density and electron-hole separation is discussed.

GPGPU implementation of growing neural gas: application to 3D scene reconstruction

Self-organising neural models have the ability to provide a good representation of the input space. In particular the Growing Neural Gas (GNG) is a suitable model because of its flexibility, rapid adaptation and excellent quality of representation. However, this type of learning is time-consuming, especially for high-dimensional input data. Since real applications often work under time constraints, it is necessary to adapt the learning process in order to complete it in a predefined time. This paper proposes a Graphics Processing Unit (GPU) parallel implementation of the GNG with Compute Unified Device Architecture (CUDA). In contrast to existing algorithms, the proposed GPU implementation allows the acceleration of the learning process keeping a good quality of representation. Comparative experiments using iterative, parallel and hybrid implementations are carried out to demonstrate the effectiveness of CUDA implementation. The results show that GNG learning with the proposed implementation achieves a speed-up of 6× compared with the single-threaded CPU implementation. GPU implementation has also been applied to a real application with time constraints: acceleration of 3D scene reconstruction for egomotion, in order to validate the proposal.

Three-dimensional planar object tracking with sub-pixel accuracy

Subpixel techniques are commonly used to increase the spatial resolution in tracking tasks. Object tracking with targets of known shape permits obtaining information about object position and orientation in the three-dimensional space. A proper selection of the target shape allows us to determine its position inside a plane and its angular and azimuthal orientation under certain limits. Our proposal is demonstrated both numerical and experimentally and provides an increase the accuracy of more than one order of magnitude compared to the nominal resolution of the sensor. The experiment has been performed with a high-speed camera, which simultaneously provides high spatial and temporal resolution, so it may be interesting for some applications where this kind of targets can be attached, such as vibration monitoring and structural analysis.

Examination of the solutions of the Navier-Stokes equations for a class of three-dimensional vortices,

"Contract AF 49(638)-255."

On one-dimensional channel flow in the presence of a magnetic field.

"Prepared for the Air Force Ballistic Missile Division, Headquarters Air Research and Development Command, under contract AF 04(647)-309 advanced propulsion systems."

The calculation of three-dimensional supersonic flows around spherically-capped smooth bodies and wings

"Work performed for the Air Force Flight Dynamics Laboratory...by the Aerodynamics Research Department of the Northrup Corporation, Aircraft Division."

Structural Insights into Viral Membrane Fusion Machinery via Cryo-Electron Tomography: Influenza Virus and Human Parainfluenza Virus

Thesis (Ph.D.)--University of Washington, 2016-06