Unravelling the fluorescence kinetics of light-harvesting proteins with simulated measurements.

The plant light-harvesting pigment-protein complex LHCII is the major antenna sub-unit of PSII and is generally (though not universally) accepted to play a role in photoprotective energy dissipation under high light conditions, a process known Non-Photochemical Quenching (NPQ). The underlying mechanisms of energy trapping and dissipation within LHCII are still debated. Various models have been proposed for the underlying molecular detail of NPQ, but they are often based on different interpretations of very similar transient absorption measurements of isolated complexes. Here we present a simulated measurement of the fluorescence decay kinetics of quenched LHCII aggregates to determine whether this relatively simple measurement can discriminate between different potential NPQ mechanisms. We simulate not just the underlying physics (excitation, energy migration, quenching and singlet-singlet annihilation) but also the signal detection and typical experimental data analysis. Comparing this to a selection of published fluorescence decay kinetics we find that: (1) Different proposed quenching mechanisms produce noticeably different fluorescence kinetics even at low (annihilation free) excitation density, though the degree of difference is dependent on pulse width. (2) Measured decay kinetics are consistent with most LHCII trimers becoming relatively slow excitation quenchers. A small sub-population of very fast quenchers produces kinetics which do not resemble any observed measurement. (3) It is necessary to consider at least two distinct quenching mechanisms in order to accurately reproduce experimental kinetics, supporting the idea that NPQ is not a simple binary switch.

Tissue Engineering Scaffold Material with Enhanced Cell Adhesion and Angiogenesis from Soy Protein Isolate Loaded with Bio Modulated Micro-TiO2 Prepared via Prolonged Sonication for Wound Healing Applications.

Tissue engineering is a technique that promotes healing by creating an ideal environment for endogenous cells to migrate and grow into the site of injury via a scaffold, improving regeneration and reducing the time required for in vitro cell culture. In this work, the effect of the addition of sonicated TiO2 in the soy protein isolate (SPI) matrix for tissue engineering applications was studied. In comparison to adding expensive nano TiO2, this method of incorporating sonicated TiO2 into the SPI matrix will aid in achieving improved properties at a lower cost. The effect of the addition of sonicated TiO2 on the morphological, UV transmittance, mechanical, thermal, surface energy, and hydrophilicity of SPI films was investigated. The result shows that the uniformly distributed TiO2 particles successfully blocked 95% of UV light. Scanning electron microscopy revealed a significant reduction in the TiO2 agglomerate size and homogeneous distribution of the same when sonication was applied instead of mechanical dispersion. A simultaneous increase of tensile strength (from 3.16 to 4.58 MPa) and elongation at break values (from 24.25% to 95.31%) with 0.5% TiO2 was observed. The addition of 0.25% TiO2 was found to significantly enhance the elongation at break value to 120.83%. Incorporation of micro-TiO2 particles could improve the surface roughness, surface energy, and wettability of SPI films. In vitro cell adhesion studies and in vivo subcutaneous implantation studies were performed to assess the cell growth and angiogenesis of the developed film membranes. An MTT assay showed that SPI-1%TiO2 film favored cell viability up to 118%, and in vivo subcutaneous implantation studies showed enhanced cell growth and angiogenesis for SPI-1% TiO2 films. This SPI-TiO2 film with enhanced surface properties can be used as an ideal candidate for tissue engineering applications.

Surface architecture of endospores of the Bacillus cereus/anthracis/thuringiensis family at the subnanometer scale.

Bacteria of the Bacillus cereus family form highly resistant spores, which in the case of the pathogen B. anthracis act as the agents of infection. The outermost layer, the exosporium, enveloping spores of the B. cereus family as well as a number of Clostridia, plays roles in spore adhesion, dissemination, targeting, and germination control. We have analyzed two naturally crystalline layers associated with the exosporium, one representing the "basal" layer to which the outermost spore layer ("hairy nap") is attached, and the other likely representing a subsurface ("parasporal") layer. We have used electron cryomicroscopy at a resolution of 0.8-0.6 nm and circular dichroism spectroscopic measurements to reveal a highly α-helical structure for both layers. The helices are assembled into 2D arrays of "cups" or "crowns." High-resolution atomic force microscopy of the outermost layer showed that the open ends of these cups face the external environment and the highly immunogenic collagen-like fibrils of the hairy nap (BclA) are attached to this surface. Based on our findings, we present a molecular model for the spore surface and propose how this surface can act as a semipermeable barrier and a matrix for binding of molecules involved in defense, germination control, and other interactions of the spore with the environment.

Cell wall peptidoglycan architecture in Bacillus subtilis.

The bacterial cell wall is essential for viability and shape determination. Cell wall structural dynamics allowing growth and division, while maintaining integrity is a basic problem governing the life of bacteria. The polymer peptidoglycan is the main structural component for most bacteria and is made up of glycan strands that are cross-linked by peptide side chains. Despite study and speculation over many years, peptidoglycan architecture has remained largely elusive. Here, we show that the model rod-shaped bacterium Bacillus subtilis has glycan strands up to 5 microm, longer than the cell itself and 50 times longer than previously proposed. Atomic force microscopy revealed the glycan strands to be part of a peptidoglycan architecture allowing cell growth and division. The inner surface of the cell wall has a regular macrostructure with approximately 50 nm-wide peptidoglycan cables [average 53 +/- 12 nm (n = 91)] running basically across the short axis of the cell. Cross striations with an average periodicity of 25 +/- 9 nm (n = 96) along each cable are also present. The fundamental cabling architecture is also maintained during septal development as part of cell division. We propose a coiled-coil model for peptidoglycan architecture encompassing our data and recent evidence concerning the biosynthetic machinery for this essential polymer.