Field Evaluation of Herbicides on Small Fruit, Vegetable, and Ornamental Crops, 1998

Growers generally use herbicides to efficiently produce high-quality fruit and vegetables for processing or fresh market sales. Due to the smaller acreage of these crops compared to major field crops, fewer herbicides are registered for use in fruit and vegetable crops than for field crops. Each year, new herbicides are evaluated under Arkansas growing conditions with the objective of improving the herbicide technology for the grower, processor, and ultimately the consumer. This report includes studies on the control of many of the more serious weed problems in important crops of this region, including snapbeans, spinach, southernpeas, watermelon, cantaloupe, summer squash, and grapes. In addition, the report includes information on the tolerance of selected bedding plants to some effective herbicides

Field Evaluation of Herbicides on Small Fruit, Vegetable, and Ornamental Crops, 1999

Growers generally use herbicides to efficiently produce high-quality fruit and vegetables for processing or fresh market sales. Because of the smaller acreage of these crops compared with major field crops, fewer herbicides are registered for use in fruit and vegetable crops than for field crops. Each year, new herbicides are evaluated under Arkansas growing conditions with the objective of improving the herbicide technology for the grower, processor, and ultimately the consumer. This report includes studies on the control of many of the more serious weed problems in important crops of this region, including snapbeans, spinach and other greens, southernpeas, tomatoes, and grapes

Spontaneous Isomerization of Long-Lived Proteins Provides a Molecular Mechanism for the Lysosomal Failure Observed in Alzheimer's Disease.

Proteinaceous aggregation is a well-known observable in Alzheimer's disease (AD), but failure and storage of lysosomal bodies within neurons is equally ubiquitous and actually precedes bulk accumulation of extracellular amyloid plaque. In fact, AD shares many similarities with certain lysosomal storage disorders though establishing a biochemical connection has proven difficult. Herein, we demonstrate that isomerization and epimerization, which are spontaneous chemical modifications that occur in long-lived proteins, prevent digestion by the proteases in the lysosome (namely, the cathepsins). For example, isomerization of aspartic acid into l-isoAsp prevents digestion of the N-terminal portion of Aβ by cathepsin L, one of the most aggressive lysosomal proteases. Similar results were obtained after examination of various target peptides with a full series of cathepsins, including endo-, amino-, and carboxy-peptidases. In all cases peptide fragments too long for transporter recognition or release from the lysosome persisted after treatment, providing a mechanism for eventual lysosomal storage and bridging the gap between AD and lysosomal storage disorders. Additional experiments with microglial cells confirmed that isomerization disrupts proteolysis in active lysosomes. These results are easily rationalized in terms of protease active sites, which are engineered to precisely orient the peptide backbone and cannot accommodate the backbone shift caused by isoaspartic acid or side chain dislocation resulting from epimerization. Although Aβ is known to be isomerized and epimerized in plaques present in AD brains, we further establish that the rates of modification for aspartic acid in positions 1 and 7 are fast and could accrue prior to plaque formation. Spontaneous chemistry can therefore provide modified substrates capable of inducing gradual lysosomal failure, which may play an important role in the cascade of events leading to the disrupted proteostasis, amyloid formation, and tauopathies associated with AD

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival

Directed-Backbone Dissociation Following Bond-Specific Carbon-Sulfur UVPD at 213 nm

Ultraviolet photodissociation or UVPD is an increasingly popular option for tandem-mass spectrometry experiments. UVPD can be carried out at many wavelengths, and it is important to understand how the results will be impacted by this choice. Here, we explore the utility of 213&nbsp;nm photons for initiating bond-selective fragmentation. It is found that bonds previously determined to be labile at 266&nbsp;nm, including carbon-iodine and sulfur-sulfur bonds, can also be cleaved with high selectivity at 213&nbsp;nm. In addition, many carbon-sulfur bonds that are not subject to direct dissociation at 266&nbsp;nm can be selectively fragmented at 213&nbsp;nm. This capability can be used to site-specifically create alaninyl radicals that direct backbone dissociation at the radical site, creating diagnostic d-ions. Furthermore, the additional carbon-sulfur bond fragmentation capability leads to signature triplets for fragmentation of disulfide bonds. Absorption of amide bonds can enhance dissociation of nearby labile carbon-sulfur bonds and can be used for stochastic backbone fragmentation typical of UVPD experiments at shorter wavelengths. Several potential applications of the bond-selective fragmentation chemistry observed at 213&nbsp;nm are discussed. Graphical Abstract ᅟ

Methionine and Selenomethionine as Energy Transfer Acceptors for Biomolecular Structure Elucidation in the Gas Phase

Mass spectrometry affords rapid and sensitive analysis of peptides and proteins. Coupling spectroscopy with mass spectrometry allows for the development of new methods to enhance biomolecular structure determination. Herein, we demonstrate two new energy acceptors that can be utilized for action-excitation energy transfer experiments. In the first system, C-S bonds in methionine act as energy acceptors from native chromophores, including tyrosine, tryptophan, and phenylalanine. Comparison among chromophores reveals that tyrosine transfers energy most efficiently at 266&nbsp;nm, but phenylalanine and tryptophan also transfer energy with comparable efficiencies. Overall, the C-S bond dissociation yields following energy transfer are low for methionine, which led to an investigation of selenomethionine, a common analog that is found in many naturally occurring proteins. Sulfur and selenium are chemically similar, but C-Se bonds are weaker than C-S bonds and have lower lying σ* anti-bonding orbitals. Excitation of peptides containing tyrosine and tryptophan results in efficient energy transfer to selenomethionine and abundant C-Se bond dissociation. A series of helical peptides were examined where the positions of the donor or acceptor were systematically scanned to explore the influence of distance and helix orientation on energy transfer. The distance was found to be the primary factor affecting energy transfer efficiency, suggesting that selenomethionine may be a useful acceptor for probing protein structure in the gas phase

Simplified identification of disulfide, trisulfide, and thioether pairs with 213 nm UVPD

Disulfide heterogeneity and other non-native crosslinks introduced during therapeutic antibody production and storage could have considerable negative effects on clinical efficacy, but tracking these modifications remains challenging. Analysis must also be carried out cautiously to avoid introduction of disulfide scrambling or reduction, necessitating the use of low pH digestion with less specific proteases. Herein we demonstrate that 213 nm ultraviolet photodissociation streamlines disulfide elucidation through bond-selective dissociation of sulfur-sulfur and carbon-sulfur bonds in combination with less specific backbone dissociation. Importantly, both types of fragmentation can be initiated in a single MS/MS activation stage. In addition to disulfide mapping, it is also shown that thioethers and trisulfides can be identified by characteristic fragmentation patterns. The photochemistry resulting from 213 nm excitation facilitates a simplified, two-tiered data processing approach that allows observation of all native disulfide bonds, scrambled disulfide bonds, and non-native sulfur-based linkages in a pepsin digest of Rituximab. Native disulfides represented the majority of bonds according to ion count, but the highly solvent-exposed heavy/light interchain disulfides were found to be most prone to modification. Production and storage methods that facilitate non-native links are discussed. Due to the importance of heavy and light chain connectivity for antibody structure and function, this region likely requires particular attention in terms of its influence on maintaining structural fidelity

Synthesis of New S-S and C-C Bonds by Photoinitiated Radical Recombination Reactions in the Gas Phase.

Photoinitiated radical chemistry has proven to be useful for breaking covalent bonds within many biomolecules in the gas phase. Herein, we demonstrate that radical chemistry is useful for bond synthesis in the gas phase. Single peptides containing two cysteine residues capped with propylmercaptan (PM) often form disulfide bonds following ultraviolet excitation at 266 nm and loss of both PM groups. Similarly, noncovalently bound peptide pairs where each peptide contains a single cysteine residue can be induced to form disulfide bonds. Comparison with disulfide bound species sampled directly from solution yields identical collisional activation spectra, suggesting that native disulfide bonds have been recapitulated in the gas phase syntheses. Another approach utilizing radical chemistry for covalent bond synthesis involves creation of a reactive diradical that can first abstract hydrogen from a target peptide, creating a new radical site, and then recombine the second radical with the new radical to form a covalent bond. This chemistry is illustrated with 2-(hydroxymethyl-3,5-diiodobenzoate)-18-crown-6 ether, which attaches noncovalently to protonated primary amines in peptides and proteins. Following photoactivation and crosslinking, the site of noncovalent adduct attachment can frequently be determined. The ramifications of these observations on peptide structure and noncovalent attachment of 18-crown-6-based molecules is discussed