Spectroscopic analyses of the noncovalent self-assembly of cyanines upon various nucleic acid scaffolds.
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We utilized self-assembly of cyanine chromophores to study the conformational changes in various types of nucleic acid scaffolds: single and double stranded DNA, linear or circular DNA and RNA. We identified a chromophore that became highly fluorescent after aggregating upon nucleic acids. Fluorescence from the aggregate was instantaneous after self-assembly. Temporal emission profiles displayed a biphasic trend demonstrating kinetic dependence for assembly and disassembly. Absorption spectra of the aggregate showed a red-shifted "shoulder" peak indicative of J-aggregate. Fluorescence from J-aggregates was also red-shifted. We utilized cyanine self-assembly to quantize various nucleic acids. The limits of detection and quantization for psiX174 DNA were 3 and 9 fmol, respectively. We similarly determined the sensitivity for various nucleic acids and established the optimum conditions for self-assembly. Collectively, the effects of methanol, salt, and full width at half maximum for cyanine fluorescence on DNA or carboxymethylamylose scaffolds, all suggested noncovalent, electrostatic, and hydrophobic forces were involved in supramolecular self-assembly. Our results facilitate a better understanding of supramolecular self-assembly. (+info)
Photoconversion in orange and red fluorescent proteins.
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Origin of slow relaxation following photoexcitation of W7 in myoglobin and the dynamics of its hydration layer.
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Molecular dynamics simulations are used to calculate the time-dependent Stokes shift following photoexcitation of Trp-7 (W7) in myoglobin. In agreement with experiment, a long time (approximately 60 ps) component is observed. Since the long time Stokes shift component is absent when we repeat the calculation with protein frozen at the instant of photoexcitation, we firmly establish that protein flexibility is required to observe slow Stokes shift dynamics in this case. A transition between sub-states near the middle of a 30 ns ground-state trajectory gave us an opportunity to compare solvation dynamics in two different environments. While some of the superficial features are different, we find that the underlying dynamics are shared by the two isomers. It is necessary to look beyond a decomposition of the Stokes shift into protein and water contributions and probe the underlying dynamics of protein side groups, backbone, and water dynamics to obtain a full picture of the relaxation process. We analyze water residence times, diffusion, and reorientation dynamics in the hydration layer. We find slow components in each of these quantities and critically examine their origin and how they affect the observed Stokes shift. (+info)
Manganese deficiency leads to genotype-specific changes in fluorescence induction kinetics and state transitions.
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