Heme geometry in the 10 K photoproduct from sperm whale carbonmonoxymyoglobin

We have measured the Soret band of the photoproduct obtained by complete photolysis of sperm whale carbonmonoxymyoglobin at 10 K. The experimental spectrum has been modeled with an analytical expression that takes into account the homogeneous bandwidth, the coupling of the electronic transition with both high and low frequency vibrational modes, and the effects of static conformational heterogeneity. The comparison with deoxymyoglobin at low temperature reveals three main differences. In the photoproduct, the Soret band is shifted to red. The band is less asymmetric, and an enhanced coupling to the heme vibrational mode at 674 cm⁻¹ is observed. These differences reflect incomplete relaxation of the active site after ligand dissociation. The smaller band asymmetry of the photoproduct can be explained by a smaller displacement of the iron atom from the mean porphyrin plane, in quantitative agreement with the X-ray structure analysis. The enhanced vibrational coupling is attributed to a subtle heme distortion from the planar geometry that is barely detectable in the X-ray structure.     

Photochemical transformations and bacterial utilization of high-molecular-weight dissolved organic carbon in a southern Louisiana tidal stream (Bayou Trepagnier)

UVirradiation of dissolved organic carbon (DOC) in the laboratory can producesmall, labile organic compounds utilizable by microbes, but few studies haveattempted to document this process in situ. ¹³Cnuclear magnetic resonance (NMR) was used to examine the bulk chemicalcomposition of natural and laboratory-irradiated high-molecular-weight DOC(HMW-DOC) from shaded (150 mol m^–2s^–1 average light in surface water) and open (1500mol m^–2 s^–1) field sitesoverone and a half years. ¹³C NMR revealed only small differences incarbon functional groups between laboratory irradiated and non-irradiatedHMW-DOC. However, bacterial protein productivity per cell (BPP) was enhanced innaturally irradiated samples of HMW-DOC in a field mesocosm experiment (p <0.05), suggesting that bacterial growth was enhanced by photochemicalproductionof labile DOC substrates. Absorbance characteristics such as spectral slope,absorbance at 350 nm, and the absorbance ratio 250nm/365 nm revealed that HMW-DOC was photoreactive,yetno differences in these values were found between samples irradiated with andwithout UV-B. In experiments conducted with simulated solar radiation in thelaboratory and with natural light in the field mesocosm experiment, UV-A(320–400 nm) and photosynthetically active radiation (PAR;400–700 nm) were more effective than UV-B (280–320nm) in HMW-DOC photolysis.     

Mercury inputs and outputs at a small lake in northern Minnesota

Storages and cycling of total mercury (Hg_T), methylmercury (MeHg), and Hg⁰ are described for Spring Lake, a small bog lake in the Marcell Experimental Forest in north-central Minnesota. We quantified photoredox transformations, MeHg photolysis, burial to the sediments, and internal and external loadings of Hg_T and MeHg. Atmospheric deposition was the main input of Hg_T; MeHg was supplied by a combination of atmospheric, near-shore wetland, and biotic (methylation) sources. Hg_T outputs were dominated by burial (67%), and Hg⁰ evasion accounted for 26% of Hg_T outputs. The watershed of Spring Lake is small (3.7× lake surface area), and accordingly, bog and upland runoff were minor contributors to both Hg_T and MeHg inputs. Wet deposition was ∼9% of total MeHg input, and other external inputs (runoff, sediment porewater) provided only an additional 7%, indicating that internal production of MeHg was occurring in the lake. Photolysis of MeHg, measured in the field and laboratory, removed ∼3× the lake mass of MeHg (20 mg) annually, and was the dominant sink for MeHg. Residence times of MeHg and Hg_T in the lake were 48 and 61 days, respectively, during the open-water season, compared with only 8 days for the residence time of MeHg on settling particles (seston). Photoreduction of Hg²⁺ to Hg⁰ was greater than the reverse reaction (Hg⁰ photooxidation), and the residence time of Hg⁰ in the photic zone was short (hours). Data from this study show active cycling of all the measured species of mercury (Hg_T, MeHg, and Hg⁰) and the importance of MeHg photolysis and photo-redox processes.     

维生素B12的光解研究及发酵液中维生素B12检测新方法的建立

采用HPLC检测法研究5,脱氧腺苷钴胺素和甲基基钴胺素的水溶液在不同光源及不同照度下的光解情况,结果表明随着光能量的增加光解速度加快。利用脱氧腺苷钴胺素的光解性质,探索了一种检测发酵液中维生素B12含量的新方法。将含有钴胺素的细胞破碎液完全光解后,测定光解后的产物羟基钴胺素,以此来确定维生素B12产量。此方法简便快速、重复性好,可应用于发酵生产维生素B12的各个环节。     

Structural, photolysis and biological studies of the bis(μ2-chloro)-tris(triphenylphosphine)-di-copper(I) and chloro-tris(triphenylphosphine)-copper(I) complexes. Study of copper(I)-copper(I) interactions

Direct reaction of copper(I) chloride with triphenylphosphine (tpp) in molar ratio 2:3 and 1:3, results in the formation of the [(tpp)Cu(μ₂-Cl)₂Cu(tpp)₂] (1) and {[CuCl(tpp)₃]·(CH₃CN)} (2) complexes. The complexes have been characterized by melting point, FT-IR, UV-Vis spectroscopic data and X-ray crystallography. Complex 1 is di-nuclear. Two μ₂-Cl atoms bridge two copper(I) ions with tetrahedral and trigonal geometry respectively. The short copper-copper bond distance of 2.9039(6) ? in case of 1 indicates d¹⁰-d¹⁰ interaction between metal centers. Thus, our studies were extended here in the determination of the quasi-aromaticity, which results in strong Cu-Cu interactions, using the computational method of nucleus-independent chemical shifts (NICS). The NICS calculated at the inner region of the Cu₂Cl₂P₃ core in complex 1 is shielded up to −6.05 ppm. Complex 2 is mono-nuclear where three phosphorus and one chloride atoms form a tetrahedron around the copper(I) ion. Photolysis of both complexes 1 and 2, results in the formation of triphenylphosphine oxide.The complexes 1 and 2, were tested for their in vitro cytotoxic activity against leiomyosarcoma cells (LMS) and human breast adenocarcinoma cells (MCF-7). The type of LMS cell death caused by the complexes was also evaluated by use of a flow cytometry assay. The results show that at concentration of 5 μΜ of complexes 1 and 2, 34.1% (1) and 19.6 (2)% of LMS cells undergo programmed cell death (apoptosis), while at 10 μΜ, 80.4% (1) and 65.2% (2) of LMS cells undergo apoptosis. The light sensitivity of the complex is discussed in relation with the biological activity.     

Distal Histidine Stabilizes Bound O2 and Acts as a Gate for Ligand Entry in Both Subunits of Adult Human Hemoglobin

The role of the distal histidine in regulating ligand binding to adult human hemoglobin (HbA) was re-examined systematically by preparing His(E7) to Gly, Ala, Leu, Gln, Phe, and Trp mutants of both Hb subunits. Rate constants for O₂, CO, and NO binding were measured using rapid mixing and laser photolysis experiments designed to minimize autoxidation of the unstable apolar E7 mutants. Replacing His(E7) with Gly, Ala, Leu, or Phe causes 20–500-fold increases in the rates of O₂ dissociation from either Hb subunit, demonstrating unambiguously that the native His(E7) imidazole side chain forms a strong hydrogen bond with bound O₂ in both the α and β chains (ΔG_{His(E7)H-bond} ≈ −8 kJ/mol). As the size of the E7 amino acid is increased from Gly to Phe, decreases in k_O2′, k_NO′, and calculated bimolecular rates of CO entry (k_entry′) are observed. Replacing His(E7) with Trp causes further decreases in k_O2′, k_NO′, and k_entry′ to 1–2 μm⁻¹ s⁻¹ in β subunits, whereas ligand rebinding to αTrp(E7) subunits after photolysis is markedly biphasic, with fast k_O2′, k_CO′, and k_NO′ values ≈150 μm⁻¹ s⁻¹ and slow rate constants ≈0.1 to 1 μm⁻¹ s⁻¹. Rapid bimolecular rebinding to an open α subunit conformation occurs immediately after photolysis of the αTrp(E7) mutant at high ligand concentrations. However, at equilibrium the closed αTrp(E7) side chain inhibits the rate of ligand binding >200-fold. These data suggest strongly that the E7 side chain functions as a gate for ligand entry in both HbA subunits.     

Unusual Spectral Properties of Bacteriophytochrome Agp2 Result from a Deprotonation of the Chromophore in the Red-absorbing Form Pr

Phytochromes are widely distributed photoreceptors with a bilin chromophore that undergo a typical reversible photoconversion between the two spectrally different forms, Pr and Pfr. The phytochrome Agp2 from Agrobacterium tumefaciens belongs to the group of bathy phytochromes that have a Pfr ground state as a result of the Pr to Pfr dark conversion. Agp2 has untypical spectral properties in the Pr form reminiscent of a deprotonated chromophore as confirmed by resonance Raman spectroscopy. UV/visible absorption spectroscopy showed that the pK_a is >11 in the Pfr form and ∼7.6 in the Pr form. Unlike other phytochromes, photoconversion thus results in a pK_a shift of more than 3 units. The Pr/Pfr ratio after saturating irradiation with monochromatic light is strongly pH-dependent. This is partially due to a back-reaction of the deprotonated Pr chromophore at pH 9 after photoexcitation as found by flash photolysis. The chromophore protonation and dark conversion were affected by domain swapping and site-directed mutagenesis. A replacement of the PAS or GAF domain by the respective domain of the prototypical phytochrome Agp1 resulted in a protonated Pr chromophore; the GAF domain replacement afforded an inversion of the dark conversion. A reversion was also obtained with the triple mutant N12S/Q190L/H248Q, whereas each single point mutant is characterized by decelerated Pr to Pfr dark conversion.     

The photolysis of octachlorodirhenate(III) in chloroform

Under 254 nm irradiation in chloroform, [Re₂Cl₈]²⁻ is converted cleanly to [Re₂Cl₉]⁻. The reaction is initiated primarily through light absorbed by the chloroform. The experimental rate expression is , where I₀ is the incident light intensity and f_S is the fraction of light absorbed by the solvent. The rate law is consistent with mechanisms involving either [Re₂Cl₈]⁻ or [Re₂Cl₉]²⁻ as an intermediate.