Two photon spectroscopy and microscopy of the fluorescent flavoprotein, iLOV.

LOV-domains are ubiquitous photosensory proteins that are commonly re-engineered to serve as powerful and versatile fluorescent proteins and optogenetic tools. The photoactive, flavin chromophore, however, is excited using short wavelengths of light in the blue and UV regions, which have limited penetration into biological samples and can cause photodamage. Here, we have used non-linear spectroscopy and microscopy of the fluorescent protein, iLOV, to reveal that functional variants of LOV can be activated to great effect by two non-resonant photons of lower energy, near infrared light, not only in solution but also in biological samples. The two photon cross section of iLOV has a significantly blue-shifted S0 → S1 transition compared with the one photon absorption spectrum, suggesting preferential population of excited vibronic states. It is highly likely, therefore, that the two photon absorption wavelength of engineered, LOV-based tools is tuneable. We also demonstrate for the first time two photon imaging using iLOV in human epithelial kidney cells. Consequently, two photon absorption by engineered, flavin-based bio-molecular tools can enable non-invasive activation with high depth resolution and the potential for not only improved image clarity but also enhanced spatiotemporal control for optogenetic applications.

Fluorescence spectroscopy and microscopy as tools for monitoring redox transformations of uranium in biological systems.

We report a study of redox reactions of uranium in model conditions using luminescence spectroscopy, which with its ease and wide availability has the potential to offer new insights into a bioremediation strategy of particular interest - the enzymatic reduction of UVIO22+ by bacteria such as Geobacter sulfurreducens. The inherent luminescent properties of UVIO22+ have been combined with confocal fluorescence microscopy techniques and lifetime image mapping to report directly on uranium concentration, localisation and oxidation state in cellular systems during uranium bioreduction, suggesting that localisation of uranyl species on the cell membrane surface plays an important role and that extracellular biogenic features form alongside uranyl sorbed cellular species during early stages of the bioreduction. The use of confocal microscopy in tandem with lifetime image mapping offers both improved temporal and spatial resolution (nanoseconds to microseconds and sub-micron respectively) than more conventional X-ray based techniques and offers the potential to image redox reactions occurring in situ. Together, these techniques provide an excellent and sensitive probe to assess the coordination environment of uranium during bioreduction processes that are currently being considered for remediation strategies of redox active radionuclides present in contaminated land.

Supramolecular interactions in PuO2Cl4(2-) and PuCl6(2-) complexes with protonated pyridines: synthesis, crystal structures, and Raman spectroscopy.

The synthesis, crystal structures, and Raman spectra of seven plutonium chloride compounds are presented. The materials are based upon Pu(VI)O2Cl4(2-) and Pu(IV)Cl6(2-) anions that are charge balanced by protonated pyridinium cations. The single crystal X-ray structures show a variety of donor-acceptor interactions between the plutonium perhalo anions and the cationic pyridine groups. Complementary Raman spectra show that these interactions can be probed through the symmetric vibrational mode of the plutonyl moiety. Unlike previously reported studies in similar uranyl(VI) systems, the facile redox chemistry of plutonium in aqueous solution has demonstrated the feasibility of using not only the An(VI)O2Cl4(2-) anion with approximate D4h symmetry but also the approximately Oh An(IV)Cl6(2-) anion in order to manipulate both the structure and dimensionality of such hybrid materials.

Synthesis, structures, and luminescent properties of uranyl terpyridine aromatic carboxylate coordination polymers.

Six novel uranyl terpyridine aromatic carboxylate coordination polymers, [UO(2)(C(6)H(2)O(4)S)(C(15)H(11)N(3))] (1), [UO(2)(C(6)H(2)O(4)S)(C(15)H(10)N(3)Cl)]·H(2)O (2), [UO(2)(C(8)H(4)O(4))(C(15)H(11)N(3))] (3), [UO(2)(C(8)H(4)O(4))(C(15)H(10)N(3)Cl)] (4), [UO(2)(C(12)H(6)O(4))(C(15)H(11)N(3))] (5), and [UO(2)(C(12)H(6)O(4))(C(15)H(10)N(3)Cl)] (6), were synthesized under solvothermal conditions and characterized by single-crystal and powder X-ray diffraction and luminescence and UV-vis spectroscopy. Compounds 1, 2, and 5 crystallize as molecular uranyl dimers, whereas compounds 3, 4, and 6 contain ladder motifs of uranyl centers. Fluorescence spectra of 1-4 show characteristic UO(2)(2+) emission, wherein bathochromic and hypsochromic shifts are noted as a function of organic species. In contrast, uranyl emission from 5 and 6 is quenched by the naphthalene dicarboxylic acid linker molecules.

Investigation of in situ oxalate formation from 2,3-pyrazinedicarboxylate under hydrothermal conditions using nuclear magnetic resonance spectroscopy.

We have investigated the assembly of a two-dimensional coordination polymer, Nd(2)(C(6)H(2)N(2)O(4))(2)(C(2)O(4))(H(2)O)(2), that has been prepared from the hydrothermal reaction of Nd(NO(3))(3)·6H(2)O and 2,3-pyrazinedicarboxylic acid (H(2)pzdc). In situ oxalate formation as observed in this system has been been investigated using (1)H and (13)C nuclear magnetic resonance spectroscopy, and a pathway for C(2)O(4)(2-) anion formation under hydrothermal conditions has been elucidated. The oxalate ligands found in Nd(2)(C(6)H(2)N(2)O(4))(2)(C(2)O(4))(H(2)O)(2) result from the oxidation of H(2)pzdc, which proceeds through intermediates, such as 2-pyrazinecarboxylic acid (2-pzca), 2-hydroxyacetamide, 3-amino-2-hydroxy-3-oxopropanoic acid, 2-hydroxymalonic acid, 2-oxoacetic acid (glyoxylic acid), and glycolic acid. The species are generated through a ring-opening that occurs via cleavage of the C-N bond of the pyrazine ring, followed by hydrolysis/oxidation of the resulting species.

Utilizing hydrogen bonds and halogen-halogen interactions in the design of uranyl hybrid materials.

Reactions of the uranyl cation (UO(2)(2+)) with 4-halopyridinium ions (X = Cl, Br, I) in high halide media (X' = Cl, Br) have produced six novel compounds, the structures of which have been determined by single crystal X-ray diffraction (XRD). The compounds can be divided into two categories based on the different modes of hydrogen bonding and halogen-halogen interactions present in the crystal structures, with one group showing approximately Type I halogen-halogen interactions and the second showing Type II. Presented is a discussion of the relative strengths of the interactions as a function of halogen size.

Rituximab plus short-duration chemotherapy as first-line treatment for follicular non-Hodgkin's lymphoma: a phase II trial of the minnie pearl cancer research network.

PURPOSE: To evaluate the feasibility and efficacy of rituximab with short-duration chemotherapy in the first-line treatment of patients with follicular non-Hodgkin's lymphoma (NHL). PATIENTS AND METHODS: Patients with previously untreated stage II-IV follicular NHL, grade 1 or 2, were eligible for this multicenter phase II trial. All patients received four weekly doses of rituximab (375 mg/m(2) intravenous), followed by three courses of combination chemotherapy (either cyclophosphamide, doxorubicin, vincristine, and prednisone [CHOP], or cyclophosphamide, vincristine, and prednisone [CVP]) plus rituximab. Patients were evaluated for response after completing treatment, and were then followed up at 3-month intervals. RESULTS: Between January 2000 and July 2001, 86 patients were treated. Eight-two patients (95%) completed treatment; no patient was withdrawn due to toxicity. The overall response rate was 93%, with 55% complete responses. After a median follow-up of 42 months, the 3- and 4-year actuarial progression-free survivals were 71% and 62%, respectively. Five patients (6%) died from lymphoma; the overall actuarial survival at 3 years was 95%. Grade 3/4 leukopenia occurred in 53% of patients, but only six patients (7%) had neutropenia or fever. Grade 3/4 nonhematologic toxicities were uncommon. CONCLUSION: Rituximab plus short-course chemotherapy is well tolerated as first-line treatment for patients with follicular NHL. The overall and complete response rates are similar to those reported with chemotherapy/rituximab combinations of longer duration. The actuarial progression-free survival of 62% at 4 years is encouraging, but further follow-up is necessary. Rituximab plus short-course chemotherapy may prove to be as effective as longer-duration chemotherapy and currently provides an attractive option for first-line treatment of elderly patients and others who tolerate chemotherapy poorly.