48 research outputs found
Recommended from our members
Competitive interactions between Douglas-fir and red alder seedlings : growth analysis, resource use, and physiology
Competitive interactions between Douglas-fir seedlings (Pseudotsuga menziesii), and seedlings of a nitrogen-fixing hardwood
associate, red alder (Alnus rubra), were investigated in an addition series experiment. The primary objective of the research was to identify and quantify key processes underlying intra- and interspecific competitive interactions of Douglas-fir and red alder seedlings. Seedlings of both species were planted into five monoculture densities of 1, 2, 4, 8, and 16 trees mâ»ÂČ, and into mixtures of all possible pairwise combinations of these densities. Stem diameter and height were measured monthly, and were used to calculate relative growth rates. Soil moisture depletion was monitored using the neutron scattering method. An index of light
availability was measured through fisheye photography and computer digitization. Leaf water potential served as a physiological
indicator of water stress physiology. Predictive equations were developed for estimating the affect of species densities on tree yield and growth as they changed with time. Patterns in resource use and
physiology were also modeled as functions of competitive regime. Yield, absolute growth, and relative growth rates based on an index
of stem volume of both species declined quantitatively in response to intra- and interspecific competition. The competitive effects of red alder intensified with time, as the dominant overstory species grew
taller and shaded the understory Douglas-fir. Resource availability and physiological performance were reduced quantitatively in response to the density of each species. Manipulation of both species
densities influenced productivity through the modification of canopy structure, light pentration, and soil moisture availability. This study demonstrated that species density, resources, and physiology interact
in a complex fashion to produce patterns of yield in stands of competing trees
Synthetic cells synthesize therapeutic proteins inside tumors
The existing dogma is that protein medicines need to be produced in large factories, and then injected to the patient. We propose that miniature artificial inert factories can be injected to the patient, to produce a protein of interest directly in the diseased tissue. We engineered artificial cell-like particles with an autonomous capacity to synthesize protein drugs after receiving an external signal. The protein is tuned to the patient\u27s needs based on a predetermined DNA code we incorporate inside the particles. This approach increases treatment efficiency and reduces adverse effects to healthy tissues.
We developed a new T7-S30 based cell-free protein synthesis system, which contains all the transcription and translation machines and molecules required for protein production (Krinsky et al., PloS one 2016). This system was used to prepare liposomes that act as artificial cells, capable of producing proteins autonomously in response to a physical trigger. Functional enzymes (luciferase and tyrosinase) and fluorescent proteins (GFP) were successfully produced using the new cell-free protein synthesis system and inside the particles both in vitro and in vivo. In addition, we demonstrated the therapeutic capabilities of the protein producing particles by producing Pseudomonas exotoxin A, an extremely potent protein, for treating cancer. Applying the particles on 4T1 cells (a triple-negative breast cancer cell-line) in vitro or injecting them into a 4T1-induced tumor in vivo, resulted in high cytotoxicity due to the effective production of the therapeutic protein inside the vesicles (Krinsky et al. Advanced Healthcare Materials, 2017).
Synthetic cells serve as autonomous, trigger-able, artificial particles that produces a variety of proteins. This platform has promise to address a wide range of fundamental questions associated with protein synthesis in nature, as well as applicative protein delivery needs.
Please click Additional Files below to see the full abstract
Recommended from our members
Predicting growth response of shrubs to clear-cutting and site preparation in coastal Oregon forests
Cover-projection models were developed based on algebraic difference formulations of an exponential-power function to describe shrub recovery and development patterns after clear-cutting and site preparation. We tested the effect of six treatments on shrub growth patterns by incorporating indicator variables into the rate and shape parameters of the models for salal (Gaultheria shallon Pursh), thimbleberry (Rubus parviflorus Nutt.), salmonberry (Rubus spectabilis Pursh), and all shrubs. For salal, the shape parameter included an adjustment for burning treatments that delayed maximum cover by several years as compared with unburned treatments. The rate parameter in the thimbleberry model was adjusted for burning treatments; maximum cover occurred about 2 years earlier in burned than in unburned treatments. Both rate and shape
parameters in the salmonberry model were adjusted for burning treatments; delayed establishment but increased growth rate and less salmonberry cover are characteristic of burned treatments as compared with the unburned treatments. The rate and shape parameters in the model for the shrub group included adjustments for burning treatments. Overstory removal fostered shrub development, whereas site preparation treatments slowed and curtailed it. The final cover-projection models accounted
for 68 92% of the total variation in cover, with the adjustments for burning accounting for 1.5 3.3% of the variation. The predicted growth patterns are consistent with trends in site occupancy and published autecological characteristics.Keywords: salal (Gaultheria shallon), overstory removal, clear-cutting, site preparation, salmonberry (Rubus spectabilis, shrub growth patterns, shrub development, burn treatment, cover-projection models, thimbleberry (Rubus parviflorus), shrub recover
Recommended from our members
Regenerating coastal forests in Oregon : an annotated bibliography of selected ecological literature
This annotated bibliography was compiled to provide a comprehensive list of sources on the ecological factors that affect forest regeneration. Abstracts from 494 publications are indexed by author, species, and subject, and are arranged into 4 major sections. Topics include the effects of biotic factors, abiotic factors, and stand and site history on vegetation, effects of vegetation on aerial and soil resources, effects of vegetation on stand development, vegetation management, plant succession, and response of vegetation to silvicultural activities and disturbances. The citations are predominately for coastal Douglas-fir occurring west of the Cascade Range in Oregon and Washington; however, applicable citations for associated conifers, hardwoods, shrubs, and herbs and for Alaska, California, and British Columbia are also included. Citations on forest genetics, seeds, seedling physiology, and insects are not included in the bibliography
Protein Engineering of Toluene Monooxygenases for Synthesis of Chiral Sulfoxidesâż
Enantiopure sulfoxides are valuable asymmetric starting materials and are important chiral auxiliaries in organic synthesis. Toluene monooxygenases (TMOs) have been shown previously to catalyze regioselective hydroxylation of substituted benzenes and phenols. Here we show that TMOs are also capable of performing enantioselective oxidation reactions of aromatic sulfides. Mutagenesis of position V106 in the α-hydroxylase subunit of toluene ortho-monooxygenase (TOM) of Burkholderia cepacia G4 and the analogous position I100 in toluene 4-monooxygenase (T4MO) of Pseudomonas mendocina KR1 improved both rate and enantioselectivity. Variant TomA3 V106M of TOM oxidized methyl phenyl sulfide to the corresponding sulfoxide at a rate of 3.0 nmol/min/mg protein compared with 1.6 for the wild-type enzyme, and the enantiomeric excess (pro-S) increased from 51% for the wild type to 88% for this mutant. Similarly, T4MO variant TmoA I100G increased the wild-type oxidation rate by 1.7-fold, and the enantiomeric excess rose from 86% to 98% (pro-S). Both wild-type enzymes showed lower activity with methyl para-tolyl sulfide as a substrate, but the improvement in the activity and enantioselectivity of the mutants was more dramatic. For example, T4MO variant TmoA I100G oxidized methyl para-tolyl sulfide 11 times faster than the wild type did and changed the selectivity from 41% pro-R to 77% pro-S. A correlation between regioselectivity and enantioselectivity was shown for TMOs studied in this work. Using in silico homology modeling, it is shown that residue I100 in T4MO aids in steering the substrate into the active site at the end of the long entrance channel. It is further hypothesized that the main function of V106 in TOM is the proper positioning or docking of the substrate with respect to the diiron atoms. The results from this work suggest that when the substrate is not aligned correctly in the active site, the oxidation rate is decreased and enantioselectivity is impaired, resulting in products with both chiral configurations
Rapid Methods for High-Throughput Detection of Sulfoxidesâż
Enantiopure sulfoxides are prevalent in drugs and are useful chiral auxiliaries in organic synthesis. The biocatalytic enantioselective oxidation of prochiral sulfides is a direct and economical approach for the synthesis of optically pure sulfoxides. The selection of suitable biocatalysts requires rapid and reliable high-throughput screening methods. Here we present four different methods for detecting sulfoxides produced via whole-cell biocatalysis, three of which were exploited for high-throughput screening. Fluorescence detection based on the acid activation of omeprazole was utilized for high-throughput screening of mutant libraries of toluene monooxygenases, but no active variants have been discovered yet. The second method is based on the reduction of sulfoxides to sulfides, with the coupled release and measurement of iodine. The availability of solvent-resistant microtiter plates enabled us to modify the method to a high-throughput format. The third method, selective inhibition of horse liver alcohol dehydrogenase, was used to rapidly screen highly active and/or enantioselective variants at position V106 of toluene ortho-monooxygenase in a saturation mutagenesis library, using methyl-p-tolyl sulfide as the substrate. A success rate of 89% (i.e., 11% false positives) was obtained, and two new mutants were selected. The fourth method is based on the colorimetric detection of adrenochrome, a back-titration procedure which measures the concentration of the periodate-sensitive sulfide. Due to low sensitivity during whole-cell screening, this method was found to be useful only for determining the presence or absence of sulfoxide in the reaction. The methods described in the present work are simple and inexpensive and do not require special equipment