Photophysical properties of 2-Phenylindole in poly (vinyl alcohol) film at room temperature. Enhanced phosphorescence anisotropy with direct triplet state excitation.

We report the spectral properties of 2-Phenylindole (2PI) embedded in rigid poly (vinyl alcohol) (PVA) film. The 2PI in PVA film shows relatively strong and structured fluorescence with a maximum at 370 nm and surprisingly strong room temperature phosphorescence with an emission maximum of about 500 nm. The dye is highly immobilized in the polymer matrix, thus presenting high fluorescence anisotropy in an isotropic film of about 0.3 at room temperature. The 2-Phenylindole phosphorescence excited in the usual way through the electronic singlet state excitation (S0 â†’ S1 absorption) results in a very low, near zero anisotropy. We now report that we can directly excite the dye to the triplet state T1 and observe high phosphorescence anisotropy similar to the fluorescence anisotropy. The extinction coefficient for S0 â†’ T1 absorption in the PVA matrix is unusually high- only about 3 orders of magnitude lower than S0 â†’ S1 absorption. We consider this direct excitation to indole's triplet state a very significant finding that may lead to many practical applications. The unusually long-wavelength of excitation around 400 nm, much above typical UV absorption, results in a high phosphorescence anisotropy. This provides a new way to study rotational motion of larger biological objects in the microsecond time scale not accessible through typical fluorescence studies.

Effect of a myosin regulatory light chain mutation K104E on actin-myosin interactions.

Familial hypertrophic cardiomyopathy (FHC) is the most common cause of sudden cardiac death in young individuals. Molecular mechanisms underlying this disorder are largely unknown; this study aims at revealing how disruptions in actin-myosin interactions can play a role in this disorder. Cross-bridge (XB) kinetics and the degree of order were examined in contracting myofibrils from the ex vivo left ventricles of transgenic (Tg) mice expressing FHC regulatory light chain (RLC) mutation K104E. Because the degree of order and the kinetics are best studied when an individual XB makes a significant contribution to the overall signal, the number of observed XBs in an ex vivo ventricle was minimized to ∼20. Autofluorescence and photobleaching were minimized by labeling the myosin lever arm with a relatively long-lived red-emitting dye containing a chromophore system encapsulated in a cyclic macromolecule. Mutated XBs were significantly better ordered during steady-state contraction and during rigor, but the mutation had no effect on the degree of order in relaxed myofibrils. The K104E mutation increased the rate of XB binding to thin filaments and the rate of execution of the power stroke. The stopped-flow experiments revealed a significantly faster observed dissociation rate in Tg-K104E vs. Tg-wild-type (WT) myosin and a smaller second-order ATP-binding rate for the K104E compared with WT myosin. Collectively, our data indicate that the mutation-induced changes in the interaction of myosin with actin during the contraction-relaxation cycle may contribute to altered contractility and the development of FHC.

The spatial distribution of actin and mechanical cycle of myosin are different in right and left ventricles of healthy mouse hearts.

The contraction of the right ventricle (RV) expels blood into the pulmonary circulation, and the contraction of the left ventricle (LV) pumps blood into the systemic circulation through the aorta. The respective afterloads imposed on the LV and RV by aortic and pulmonary artery pressures create very different mechanical requirements for the two ventricles. Indeed, differences have been observed in the contractile performance between left and right ventricular myocytes in dilated cardiomyopathy, in congestive heart failure, and in energy usage and speed of contraction at light loads in healthy hearts. In spite of these functional differences, it is commonly believed that the right and left ventricular muscles are identical because there were no differences in stress development, twitch duration, work performance, or power among the RV and LV in dogs. This report shows that on a mesoscopic scale [when only a few molecules are studied (here three to six molecules of actin) in ex vivo ventricular myofibrils], the two ventricles in rigor differ in the degree of orientational disorder of actin within in filaments and during contraction in the kinetics of the cross-bridge cycle.

Long lived BSA Au clusters as a time gated intensity imaging probe.

The work presented here reports the use of long lifetime (>1 µs) BSA Au clusters as a cellular/tissue, time gated, intensity imaging probe. By collecting the emission signal 50 ns post excitation, one can off-gate the intense auto-fluorescence background, thereby greatly enhancing the clarity/specificity in fluorescence imaging.

New structures of the O-specific polysaccharides of proteus. 4. Polysaccharides containing unusual acidic N-acyl derivatives of 4-amino-4,6-dideoxy-D-glucose.

The structures of the O-polysaccharides of the lipopolysaccharides of Proteus mirabilis O7 and O49 were determined by chemical methods, mass spectrometry, including MS/MS, and NMR spectroscopy, including experiments run in an H2O/D2O mixture to reveal correlations for NH protons. The O-polysaccharides were found to contain N-carboxyacetyl (malonyl) and N-(3-carboxypropanoyl) (succinyl) derivatives of 4-amino-4,6-dideoxyglucose (4-amino-4-deoxyquinovose, Qui4N), respectively. The behavior of Qui4N derivatives with the dicarboxylic acids under conditions of acid hydrolysis and methanolysis was studied using GLC-MS.