Antidiabetic Potential of Protein Hydrolysates and Peptide Fractions from Lima Bean (Phaseolus lunatus L): An In Vitro Study

International Journal of Peptide Research and Therapeutics - Characterized by uncontrolled, long-term high blood sugar levels, diabetes mellitus affects ever increasing numbers of people worldwide....     

Passive Internalization of Bioactive β-Casein Peptides into Phospholipid (POPC) Bilayers. Free Energy Landscapes from Unbiased Equilibrium MD Simulations at μs-Time Scale

Food Biophysics - Absorption of bioactive peptides in the intestinal epithelium take place in the apical or the basolateral tight junctions of the cells. Depending on the peptide size and...     

Conformational insights on the molecular recognition processes of carbohydrate molecules by proteins and enzymes: A 3D view by using NMR

This review focuses, in a non-exhaustive manner, on the essential structural and conformational features of protein–carbohydrate interactions and on some applications of NMR spectroscopy to deal with this topic from different levels of complexity.     

Long-term biomonitoring of lichen and bryophyte biodiversity at Burnham Beeches SAC and global environmental change

Long-term monitoring began 20 years ago at Burnham Beeches Site of Special Interest (SSSI), National Nature Reserve (NNR) and European Special Area of Conservation (SAC) lying 40 km west of London as a consequence of the authorization of an application to extract gravel from an adjacent site lying north of Slough Trading Estate. Dust monitoring (sticky pads) and photographic monitoring, recording and image analysis was instigated in 1992 on Parmelion communities to assess changes in lichen growth, health and community composition. Long-term monitoring identifies that the lichen flora on free-standing trees has undergone rapid expansion from a near dominance by the SO₂-tolerant ‘acidophyte’ species Lecanora conizaeoides and Hypogymnia physodes following reductions in SO₂ concentrations. Long-term influences of low levels of eutrophication, gaseous pollutants (particularly globally rising background ozone concentrations) on lichen and bryophyte communities and succession under changing climatic conditions are unknown. Soil–plant relationships, lichen–invertebrate interactions and a pollution legacy must also be considered.     

The Design and Synthesis of N 4-Anthraniloyl-2′-dC,the Improved Syntheses of N 4-Carbamoyl-and N4- Ureidocarbamoyl-2′-dC,Incorporation into Oligonucleotides and Triplex Formation Testing

Abstract

Three modified nucleosides were designed with the aim of achieving triplet formation with the CG base pair of duplex DNA. Direct anthraniloylation of 2′-deoxycytidine, using isatoic anhydride, afforded the novel N ⁴-anthraniloyl-2′-deoxycytidine. Much improved preparations of N ⁴-carbamoyl-2′-deoxycytidine and of N ⁴-ureidocarbonyl-2′-deoxycytidine were accomplished. The modified nucleosides were incorporated into oligonucleotides. Thermal denaturation studies and gel mobility shift analysis suggest that these nucleosides do not form base triplets with any of the four base pairs of DNA.     

Identification and Quantification of AKT Isoforms and Phosphoforms in Breast Cancer Using a Novel Nanofluidic Immunoassay

Breast cancer subtype-specific molecular variations can dramatically affect patient responses to existing therapies. It is thought that differentially phosphorylated protein isoforms might be a useful prognostic biomarker of drug response in the clinic. However, the accurate detection and quantitative analysis of cancer-related protein isoforms and phospho-isoforms in tumors are limited by current technologies. Using a novel, fully automated nanocapillary electrophoresis immunoassay (NanoPro^TM 1000) designed to separate protein molecules based on their isoelectric point, we developed a reliable and highly sensitive assay for the detection and quantitation of AKT isoforms and phosphoforms in breast cancer. This assay enabled the measurement of activated AKT1/2/3 in breast cancer cells using protein produced from as few as 56 cells. Importantly, we were able to assign an identity for the phosphorylated S473 phosphoform of AKT1, the major form of activated AKT involved in multiple cancers, including breast, and a current focus in clinical trials for targeted intervention. The ability of our AKT assay to detect and measure AKT phosphorylation from very low amounts of total protein will allow the accurate evaluation of patient response to drugs targeting activated PI3K-AKT using scarce clinical specimens. Moreover, the capacity of this assay to detect and measure all three AKT isoforms using one single pan-specific antibody enables the study of the multiple and variable roles that these isoforms play in AKT tumorigenesis.Activation of the PI3K-AKT signaling pathway is one of the most common events in cancer (1, 2). Pathway activation can confer a number of advantages to the cancer cells, including enhanced proliferation and survival (1, 2). Multiple mechanisms exist by which the pathway may become activated, including amplification or activation of receptor tyrosine kinases (e.g. ERBB2 in breast and EGFR in lung tumors), mutation of the catalytic or regulatory subunits of PI3K (e.g. PIK3CA in colorectal and breast tumors), loss of the negative regulator PTEN (e.g. mutation in prostate and melanoma), and gain of function of AKT (e.g. amplification or mutation in breast and pancreatic tumors) (reviewed in Refs. 1 and 2).AKT represents a central node in the PI3K signaling cascade (3). AKT is recruited to the cell membrane via its pleckstrin homology domain when PI3K phosphorylates PIP₂ to form PIP₃ (4, 5). Following recruitment, AKT is phosphorylated by PDK1 and the rictor-mTOR complex, resulting in conformational changes and activation of the protein (5–8). Multiple studies have shown that the phosphorylation of AKT leads to the phosphorylation and activation of downstream effectors of the signaling pathway, such as mTOR complex 1 and S6K (reviewed in Ref. 1). The central role of this pathway in cancer is further underscored by the efforts of multiple pharmaceutical companies that have developed inhibitors against AKT as potential anti-oncogenic therapeutics (9).Despite the importance of AKT in growth and survival signaling in cancer, there are surprisingly few data that address the specific roles played in growth and survival by the multiple AKT family members (AKT-1, -2, and -3) and different phosphorylation and putative phosphorylation sites that can potentially activate the protein. Western blot analysis has been the foundation of most AKT studies, but in many cases pan-AKT antibodies have been employed that fail to distinguish between the different AKT isoforms. Recent siRNA silencing studies have indicated distinct functions for different AKT family members within a cell (10, 11). Moreover, there is evidence in breast cancer that the three isoforms exhibit different localizations and therefore must have at least partially distinct functions (12). Similarly, evidence is mounting for multiple phosphorylation sites in AKT beyond the two most studied phosphorylation events (Thr-308 and Ser-473) (5–8). Phosphorylation at serine and threonine residues at Thr-72 and Ser-246 may be required for the activation or regulation of kinase activity (13). The functional significance of constitutive phosphorylation of Ser-124 and Thr-450 is still unknown (14). Finally, there is evidence that phosphorylation of tyrosine residues at Tyr-315 and Tyr-326 is required for full kinase activity (15).Analysis of such phospho- and isoform-specific activation often requires complicated in-depth analyses using large quantities of proteins, purified recombinant protein, immunoprecipitation, incorporation of ³²P isotopes, and/or mass spectroscopy, which makes such studies more difficult to perform and not easily adaptable to clinical specimens. Thus, better methods are required for the accurate assessment of both phosphoform and isoform usage in cells with an activated PI3K-AKT pathway and the effects of pathway inhibitors using relatively small amounts of starting material. We describe here the development of such an assay using nanocapillary-based isoelectric focusing (16). This approach allows the separation of AKT into distinct peaks that correspond to different iso- and phosphoforms using a small amount of starting material and a single pan-specific antibody. This approach should allow for more accurate determinations of isoform usage in different cell types, as well as of changes in phosphorylation states in response to pathway inhibition, including in clinical specimens.     

Beta‐diversity in temperate and tropical forests reflects dissimilar mechanisms of community assembly

Site‐to‐site variation in species composition (β‐diversity) generally increases from low‐ to high‐diversity regions. Although biogeographical differences in community assembly mechanisms may explain this pattern, random sampling effects can create this pattern through differences in regional species pools. Here, we compared assembly mechanisms between spatially extensive networks of temperate and tropical forest plots with highly divergent species pools (46 vs. 607 species). After controlling for sampling effects, β‐diversity of woody plants was similar and higher than expected by chance in both forests, reflecting strong intraspecific aggregation. However, different mechanisms appeared to explain aggregation in the two forests. In the temperate forest, aggregation reflected stronger environmental correlations, suggesting an important role for species‐sorting (e.g. environmental filtering) processes, whereas in the tropics, aggregation reflected stronger spatial correlations, more likely reflecting dispersal limitation. We suggest that biogeographical differences in the relative importance of different community assembly mechanisms contribute to these striking gradients in global biodiversity.