Langmuir-Blodgett (LB) technology was used to build a high-sensitivity enzyme-based biosensor for medical purposes. Recombinant fungal laccase from Rigidoporous lignosus, as previously described, was used to catalyze a widely used antidepressant in a micromolar range, namely, clomipramine. The topological properties of the laccase thin film were characterized via LB π-A isotherm and AFM (mean roughness 8.22 nm, compressibility coefficient 37.5 m/N). The sensitivity of the biosensor was investigated via UV spectroscopy, and linearity was found in the absorbance peak shift at 400 nm at drug concentration varying up to 20 uM. The enzyme kinetics was subsequently investigated with potentiometric and amperometric measurements, and we found electronic transfer of at least 1 electron, k(s) 0.57 s(-1), diffusion coefficient 3 × 10(-6) cm(2)/s, K(cat) 6825.92 min(-1), K(M) 4.1 uM, K(cat)/K(M) 2.8 × 10(7) mol(-1) s(-1), sensitivity of 440 nA/uM, maximum velocity 1706.48 nA/s, and response time less than 5 s. The amperometric and potentiometric measurements were repeated after a month, confirming the stability of the biosensor.
X-ray atomic structure of recombinant Hell's gate globin I (HGbI) from Methylacidophilum infernorum was calculated from the X-ray diffraction data of two different types of crystals: obtained by classical hanging drop and by LB nanotemplate method under the same crystallization conditions. After the accurate comparison of crystallographic parameters and electron density maps of two structures they appears to be quite similar, while the quality of the crystals grown by LB nanotemplate method was higher then of those grown by classical method. Indeed, the resolution of the LB crystal structure was 1.65 Å, while classical crystals showed only 3.2 Å resolution. Moreover, the reproducibility of this result in the case of LB crystals was much better-nine crystals from 10 gave the same structural results, while only two of 10 classical crystals were appropriate for the X-ray structure resolution.
A state-of-the-art review of the role of the Langmuir-Blodgett nanotemplate on protein crystal structures is here presented. Crystals grown by nanostructured template appear more radiation resistant than the classical ones, even in the presence of a third-generation highly focused beam at the European Synchrotron Radiation Facility. The electron density maps and the changes in parameters such as total diffractive power, B-factor, and pairwise R-factor have been discussed. Protein crystals, grown by the Langmuir-Blodgett nanotemplate-based method, proved to be more radiation resistant compared to crystals grown by the classical hanging drop method in terms of both global and specific damage.
Matrices and sensors resulting from inorganic, organic and biological nanocomposites are presented in this overview. The term nanocomposite designates a solid combination of a matrix and of nanodimensional phases differing in properties from the matrix due to dissimilarities in structure and chemistry. The nanoocomposites chosen for a wide variety of health and environment sensors consist of Anodic Porous Allumina and P450scc, Carbon nanotubes and Conductive Polymers, Langmuir Blodgett Films of Lipases, Laccases, Cytochromes and Rhodopsins, Three-dimensional Nanoporous Materials and Nucleic Acid Programmable Protein Arrays.
Sympatry of amphimictic and parthenogenetic lineages in species with mixed reproductive systems is rarely observed in nature. On Lampedusa Island (Pelagie Islands, Italy), amphimictic and parthenogenetic lineages of Heterocypris barbara (Gauthier and Brehm, 1928) co-occur in a temporary pond. Their sympatric persistence calls for an ecological differentiation. We investigated the role of seasonal variation of temperature and photoperiod conditions by two different approaches: microcosms set up by inundation of dry sediments from the temporary pond and life-table experiments. Microcosms recreate conditions similar to the field and in their sediments random samples of resting eggs of both amphimictic and parthenogenetic females are stored. Life-table experiments supplied individual-based estimates of survivorship, adult life span, fecundity, and sex ratio in the progeny. We carried out the experiments at 24 °C and a photoperiod of 12 h light (L) : 12 h dark (D) (simulating fall conditions) and at 16 °C and a photoperiod of 10 h L : 14 h D (simulating winter conditions). Males and amphimictic females were the most numerous forms at 24 °C and 12 h L : 12 h D; parthenogenetic females were dominant at 16 °C and 10 h L : 14 h D. Life-table experiments showed that amphimictic forms do not complete development at 16 °C and 10 h L : 14 h D. Our results suggest that sympatry of amphimictic and parthenogenetic females in the field depends on seasonal niche partitioning and the storage effect of resting eggs that allows survival through adverse-season conditions.
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