Actuated plasmonic nanohole arrays for sensing and optical spectroscopy applications.

Herein, we report a new approach to rapidly actuate the plasmonic characteristics of thin gold films perforated with nanohole arrays that are coupled with arrays of gold nanoparticles. The near-field interaction between the localized and propagating surface plasmon modes supported by the structure was actively modulated by changing the distance between the nanoholes and nanoparticles and varying the refractive index symmetry of the structure. This approach was applied by using a thin responsive hydrogel cushion, which swelled and collapsed by a temperature stimulus. The detailed experimental study of the changes and interplay of localized and propagating surface plasmons was complemented by numerical simulations. We demonstrate that the interrogation and excitation of the optical resonance to these modes allow the label-free SPR observation of the binding of biomolecules, and is applicable for in situ SERS studies of low molecular weight molecules attached in the gap between the nanoholes and nanoparticles.

Compact Grating-Coupled Biosensor for the Analysis of Thrombin.

A compact optical biosensor for direct detection of thrombin in human blood plasma (HBP) is reported. This biosensor platform is based on wavelength spectroscopy of diffraction-coupled surface plasmons on a chip with a periodically corrugated gold film that carries an antifouling thin polymer layer consisting of poly[(N-(2-hydroxypropyl)methacrylamide)-co-(carboxybetaine methacrylamide)] (poly(HPMA-co-CBMAA)) brushes. This surface architecture provides superior resistance to nonspecific and irreversible adsorption of abundant compounds in the analyzed HBP samples in comparison to standard surface modifications. The carboxylate groups along the polymer brushes were exploited for the covalent immobilization of aptamer ligands. These ligands were selected to specifically capture the target thrombin analyte from the analyzed HBP sample in a way that does not activate the coagulatory process at the biosensor surface with poly(HPMA-co-CBMAA) brushes. Direct label-free analysis of thrombin in the medically relevant concentration range (1-20 nM) is demonstrated without the need for diluting the HBP samples or using additional steps for signal enhancement. The reported platform constitutes the first step toward a portable and sensitive point-of-care device for direct detection of thrombin in human blood.

Correction: New Insight into Metal Ion-Driven Catalysis of Nucleic Acids by Influenza PA-Nter.

[This corrects the article DOI: 10.1371/journal.pone.0156972.].

New Insight into Metal Ion-Driven Catalysis of Nucleic Acids by Influenza PA-Nter.

PA subunit of influenza RNA-dependent RNA polymerase deserves constantly increasing attention due to its essential role in influenza life cycle. N-terminal domain of PA (PA-Nter) harbors endonuclease activity, which is indispensable in viral transcription and replication. Interestingly, existing literature reports on in vitro ion preferences of the enzyme are contradictory. Some show PA-Nter activity exclusively with Mn2+, whereas others report Mg2+ as a natural cofactor. To clarify it, we performed a series of experiments with varied ion concentrations and substrate type. We observed cleavage in the presence of both ions, with a slight preference for manganese, however PA-Nter activity highly depended on the amount of residual, co-purified ions. Furthermore, to quantify cleavage reaction rate, we applied fluorescence cross-correlation spectroscopy (FCCS), providing highly sensitive and real-time monitoring of single molecules. Using nanomolar ssDNA in the regime of enzyme excess, we estimated the maximum reaction rate at 0.81± 0.38 and 1.38± 0.34 nM/min for Mg2+ and Mn2+, respectively. However, our calculations of PA-Nter ion occupancy, based on thermodynamic data, suggest Mg2+ to be a canonical metal in PA-Nter processing of RNA in vivo. Presented studies constitute a step toward better understanding of PA-Nter ion-dependent activity, which will possibly contribute to new successful inhibitor design in the future.

[AMP-activated protein kinase (AMPK) as therapeutic target]. / Kinaza bialkowa aktywowana przez AMP (AMPK) jako cel terapeutyczny.

AMP-activated protein kinase (AMPK) is one of the major energy sensor at both: cellular and whole body level. It exists as heterotrimer containing three subunits: the catalytic α subunit, ß and regulatory γ. AMPK is localized both in the cytoplasm and in the nucleus. It is activated by increasing concentrations of AMP during the energy shortage, causing activation of catabolic pathways and inhibition of energy consuming processes. AMPK activity can be regulated allosterically: by binding AMP to a regulatory γ subunit, as well as by phosphorylation on Thr172 of the catalytic α subunit by other kinases. Activated AMPK can effectively inhibit the mTOR pathway which is hyperactive in many types of cancer. On the other hand AMPK inactivation associates with the type II diabetes, diet-induced obesity, insulin resistance and the development of other metabolic disorders. The AMPK dysfunction is also observed in inflammation. It was discovered during last years that abnormalities in the AMPK function can induce the metabolic reprogramming in cancer cells known as the Warburg effect. Additionally, AMPK is activated during irradiation. Its activation leads to inhibition of growth. On the other hand, active AMPK enables cells to survive in difficult conditions such as hypoxia, or glucose deprivation. Because of its crucial role in maintaining of the energy homeostasis AMPK is an excellent therapeutic target. However, it still remains unknown what is better: to activate or inhibit the AMPK function.