Electrostatic and conformational effects on the electronic structures of distortional isomers of a mixed-valence binuclear Cu complex.

The electronic structure of the binuclear copper complex [Cu(2)(L)](3+) [L = N(CH(2)CH(2)N(H)CH(2)CH(2)N(H)CH(2)CH(2))(3)N] has been investigated by resonance Raman and electroabsorption spectroscopy. Crystallographic Cu(2) distances of 2.364(1) and 2.415(1) A determined for the nitrate and acetate salts, respectively, are consistent with a substantial metal-metal interaction. The Cu-Cu bonding interaction in the binuclear complex is modulated both in the solid state and in solution by the ligand environment through coupling to ligand torsional modes that are, in turn, stabilized by hydrogen bonding. Electroabsorption data on the three major visible and near-infrared electronic transitions of Cu(2)L, lambda(max) (epsilon(max)) = 1000 nm ( approximately 1200 M(-1) cm(-1)), 748 nm (5600 M(-1) cm(-1)), and 622 nm (3350 M(-1) cm(-1)), reveal a difference dipole moment between the ground and excited states (Deltamu(A)) because of symmetry breaking. The difference polarizability for all three of the transitions is negative, indicating that the ground state is more polarizable than the excited state. A general model to explain this behavior in terms of the proximity of accessible transitions involving copper d electrons is proposed to explain the larger polarizability of the ground state. Raman excitation profiles (REPs) provide evidence for multiple conformational states of [Cu(2)(L)](3+). Separate REPs were obtained for each of the components of the two major Raman bands for nu(1) (a Cu-Cu stretching mode) and nu(2) (a Cu-Cu-N(eq) bending mode). The Raman data along with quantum chemical ZINDO/S CI calculations provide evidence for isomeric forms of Cu(2)L with strong coupling between the conformation of L and the Cu-Cu bond length.

Hypertonic saline infusion: can it regulate human neutrophil function?

The down-regulation of neutrophil adhesion molecule expression after hemorrhagic shock may reduce neutrophil-mediated organ injury. Hypertonic saline (HS) blocks neutrophil activation, and HS infusion in animals reduces organ injury. In this study, we investigated whether HS infusion in healthy human volunteers can affect neutrophil function. Healthy human volunteers were administered either 4 mL/kg of a 7.5% HS (n = 6) or normal saline (NS, 0.9%; n = 5) over 15 min. Mean arterial pressure (MAP) and plasma sodium levels were measured. Blood samples were obtained before and 1 h after fluid administration. Cells were stimulated with fMLP or left untreated. Neutrophil phagocytosis and expression of CD11b and L-selectin was determined with flow cytometry. HS infusion caused a 7 +/- 2 mM rise in plasma Na+ levels that was sustained at 6 +/- 1 mM for 60 min. MAP was affected only in one subject. HS and NS infusion had little effect on neutrophil phagocytosis. After HS infusion, CD1lb expression of unstimulated neutrophils was 26 +/- 6% lower than before HS infusion, and that of fMLP-stimulated cells was 12 +/- 2% lower compared to pre-infusion values. NS infusion had no significant effects on neutrophil CD11b expression. L-selectin expression of unstimulated cells after HS infusion was 9 +/- 3% higher than in the pre-infusion samples. These data suggest that HS infusion could indeed affect human neutrophils by suppressing CD11b expression. Although modest in healthy subjects, this effect may be more pronounced in trauma patients where reduced neutrophil-endothelial cell interactions might lessen neutrophil-mediated tissue damage.

Temperature dependence of the Qy resonance Raman spectra of bacteriochlorophylls, the primary electron donor, and bacteriopheophytins in the bacterial photosynthetic reaction center.

Qy-excited resonance Raman spectra of the accessory bacteriochlorophylls (B), the bacteriopheophytins (H), and the primary electron donor (P) in the bacterial photosynthetic reaction center (RC) of Rhodobacter sphaeroides have been obtained at 95 and 278 K. Frequency and intensity differences are observed in the low-frequency region of the P vibrational spectrum when the sample is cooled from 278 to 95 K. The B and H spectra exhibit minimal changes of frequencies and relative intensities as a function of temperature. The mode patterns in the Raman spectra of B and H differ very little from Raman spectra of the chromophores in vitro. The Raman scattering cross sections of B and H are 6-7 times larger than those for analogous modes of P at 278 K. The cross sections of B and of H are 3-4 times larger at 95 K than at 278 K, while the cross sections of P are approximately constant with temperature. The temperature dependence of the Raman cross sections for B and H suggests that pure dephasing arising from coupling to low-frequency solvent/protein modes is important in the damping of their excited states. The weak Raman cross sections of the special pair suggest that the excited state of P is damped by very rapid (<<30 fs) electronic relaxation processes. These resonance Raman spectra provide information for developing multimode vibronic models of the excited-state structure and dynamics of the chromophores in the RC.

Heterologous expression of an engineered biosynthetic pathway: functional dissection of type II polyketide synthase components in Streptomyces species.

Polyketides are an extensive class of secondary metabolites with diverse molecular structures and biological activities. A plasmid-based multicomponent polyketide synthase expression cassette was constructed using a subset of actinorhodin (act) biosynthetic genes (actI-orf1, actI-orf2, actI-orf3, actIII, actVII, and actIV) from Streptomyces coelicolor which specify the construction of the anthraquinone product aloesaponarin II, a molecule derived from acetyl coenzyme A and 7 malonyl coenzyme A extender units. This system was designed as an indicator pathway in Streptomyces parvulus to quantify polyketide product formation and to examine the functional significance of specific polyketide synthase components, including the act beta-ketoacyl synthase (beta-KS; encoded by actI-orf1 and actI-orf2) and the act cyclase/dehydrase (encoded by actVII-orf4). Site-directed mutagenesis of the putative active site Cys (to a Gln) in the actI-orf1 beta-KS product completely abrogated aloesaponarin II production. Changing the putative acyltransferase active-site Ser (to a Leu) located in the actI-orf1 beta-KS product led to significantly reduced but continued production of aloesaponarin II. Replacement of the expression cassette with one containing a mutant form of actI-orf2 gave no production of aloesaponarin II or any other detectable polyketide products. However, an expression cassette containing a mutant form of actVII-orf4 gave primarily mutactin with low-level production of aloesaponarin II.

Rapid-flow resonance Raman spectroscopy of bacterial photosynthetic reaction centers.

Rapid-flow resonance Raman vibrational spectra of bacterial photosynthetic reaction centers from the R-26 mutant of Rhodobacter sphaeroides have been obtained by using excitation wavelengths (810-910 nm) resonant with the lowest energy, photochemically active electronic absorption. The technique of shifted excitation Raman difference spectroscopy is used to identify genuine Raman scattering bands in the presence of a large fluorescence background. The comparison of spectra obtained from untreated reaction centers and from reaction centers treated with the oxidant K3Fe(CN)6 demonstrates that resonance enhancement is obtained from the special pair. Relatively strong Raman scattering is observed for special pair vibrations with frequencies of 36, 94, 127, 202, 730, and 898 cm-1; other modes are observed at 71, 337, and 685 cm-1. Qualitative Raman excitation profiles are reported for some of the strong modes, and resonance enhancement is observed to occur throughout the near-IR absorption band of the special pair. These Raman data determine which vibrations are coupled to the optical absorption in the special pair and, thus, probe the nuclear motion that occurs after electronic excitation. Implications for the interpretation of previous hole-burning experiments and for the excited-state dynamics and photochemistry of reaction centers are discussed.

Femtosecond energy-transfer processes in the B800-850 light-harvesting complex of Rhodobacter sphaeroides 2.4.1.

The B800-to-B850 energy transfer time in the purified B800-850 light-harvesting complex of Rhodobacter sphaeroides 2.4.1 is determined to be 0.7 ps at room temperature. The electronic state dynamics of the principal carotenoid of this species, spheroidene, are examined, both in vivo and in vitro, by direct femtosecond time-resolved experiments and by fluorescence emission yield studies. Evidence is presented which suggests that carotenoid-to-bacteriochlorophyll energy transfer may occur directly from the initially excited carotenoid S2 state, as well as from the carotenoid S1 state. Further support for this conjecture is obtained from calculations of energy transfer rates from the carotenoid S2 state. Previous measurements of in vivo carotenoid and B800 dynamics are discussed in light of the new results, and currently unresolved issues are described.

Femtosecond dynamics of energy transfer in B800-850 light-harvesting complexes of Rhodobacter sphaeroides.

We report femtosecond transient absorption studies of energy transfer dynamics in the B800-850 light-harvesting complex (LHC) of Rhodobacter sphaeroides 2.4.1. For complexes solubilized in lauryldimethylamine-N-oxide (LDAO), the carotenoid to bacteriochlorophyll (Bchl) B800 and carotenoid to Bchl B850 energy transfer times are 0.34 and 0.20 ps, respectively. The B800 to B850 energy transfer time is 2.5 ps. For complexes treated with lithium dodecyl sulfate (LDS), a carotenoid to B850 energy transfer time of less than or equal to 0.2 ps is seen, and a portion of the total carotenoid population is decoupled from Bchl. In both LDAO-solubilized and LDS-treated complexes an intensity-dependent picosecond decay component of the excited B850 population is ascribed to excitation annihilation within minimal units of the LHC.

Glucose metabolism of various tissues of pear buds.

Various tissues in flower buds of Pyrus calleryana Decne. differ in their metabolic activity. Brown outer scales utilized more exogenously supplied glucose, particularly through the pentose phosphate pathway, than did the central axes and the green inner scales. They also contained more endogenous reducing sugars, and glucose leaked out more readily from the brown scales than from the other tissues. In contrast, respiration of the central axes was nine times as great as that of the brown scales, and two to four times as much glucose was metabolized through glycolysis. Membranes of the central axes were less permeable to glucose. Because the brown scales are 75% of the dry weight of the bud, they dominate its pattern of glucose metabolism.