Photochemical Colorine Reaction of Sake

Coloring reaction of sake caused by exposure to sun-light was investigated. There is one of the coloring reactions in which deferri-ferrichrysin participate. The reaction was named Reaction I.

Deferri-ferrichrysin, tyrosine, Mn²⁺ and unknown nitrogenous compound named conveniently compound X were essential for Reaction I and citric acid was stimulative.

Compound X was purified by using Amberlite IR 120 (H type) column, active carbon column, silicic acid column and alumina column, and crystallized from methanol-water (1 : 1).

The crystals decomposed at 288~290°C. The ultraviolet and infrared spectra of the compound X were essentially identical with those of authentic kynurenic acid.

From these results the compound X was identified as kynurenic acid.

Since kynurenic acid alone did not cause the color development and riboflavin could be substituted for kynurenic acid in Reaction I, kynurenic acid may act as photosensitizer in Reaction I.     

The Photochemical Reaction of Pentachlorophenol

The chemical structure of a yellow C₁₈-compound (IV), isolated from the decomposition products of sodium pentachlorophenoxide (Na-PCP) in an aqueous solution by sunlight, has been determined by chemical and spectroscopic evidences. Some of the chemistry and the absorption spectra of IV and its related compounds containing 3-cyclohexene-1,2-dione and spiroketal structures are also described.     

The Photochemical Reaction of Pentachlorophenol

A herbicide, sodium pentachlorophenoxide (Na-PCP), used in Japan, is easily decomposed with sunlight after its application in the rice field. The photochemical reaction of Na-PCP in an aqueous solution on exposure to sunlight afforded numerous products which were mainly accompanied with chloranilic acid and a yellow compound (I). The chemical structure of the yellow compound I was established as being 3, 4, 5-trichloro-6-(2′-hydroxy-3′, 4′, 5′, 6′-tetrachlorophenoxy)-o-benzoquinone by chemical and spectroscopic evidences.

Minor decomposition products of sodium pentachlorophenoxide (Na-PCP) in an aqueous solution by sunlight have been isolated. Chemical structures of them are described, and infrared and ultraviolet spectra are presented in support of these stractures. These illustrate a new type of oxidative reaction of phenols.     

Photochemical Reaction between Uridine and Sulfhydryl Compounds

Kinetics of photochemical reaction between uridine and sulfhydryl compounds were investigated in phosphate buffer (pH 7.0) and in unbuffered aqueous solution under aerobic condition. The results obtained clearly demonstrate that the photo-induced hydration or reduction of the uridine molecule was significantly influenced by the amount of sulfhydryl group present in the reaction medium. Reaction on uridine (1 mm) was observed to lead to photohydration with pseudo first order rates, independently of the presence of cysteine (1 mm or 2 mm), while in the presence of dithiothreitol (DTT, 1 mm to 10 mm) both photoreduction and photohydration of uridine were observed. The rate of photoreduction came to predominate as the amount of DTT increased. The reaction was discussed from the view point of food chemistry as well as reaction pathways.     

The Photochemical Reaction Cycle and Photoinduced Proton Transfer of Sensory Rhodopsin II (Phoborhodopsin) from Halobacterium salinarum

Sensory rhodopsin II (HsSRII, also called phoborhodopsin) is a negative phototaxis receptor of Halobacterium salinarum, a bacterium that avoids blue-green light. In this study, we expressed the protein in Escherichia coli cells, and reconstituted the purified protein with phosphatidylcholine. The reconstituted HsSRII was stable. We examined the photocycle by flash-photolysis spectroscopy in the time range of milliseconds to seconds, and measured proton uptake/release using a transparent indium-tin oxide electrode. The pKa of the counterion of the Schiff base, Asp⁷³, was 3.0. Below pH 3, the depleted band was observed on flash illumination, but the positive band in the difference spectra was not found. Above pH 3, the basic photocycle was HsSRII (490) → M (350) → O (520) → Y (490) → HsSRII, where the numbers in parentheses are the maximum wavelengths. The decay rate of O-intermediate and Y-intermediate were pH-independent, whereas the M-intermediate decay was pH-dependent. For 3 < pH < 4.5, the M-decay was one phase, and the rate decreased with an increase in pH. For 4.5 < pH < 6.5, the decay was one phase with pH-independent rates, and azide markedly accelerated the M-decay. These findings suggest the existence of a protonated amino acid residue (X-H) that may serve as a proton relay to reprotonate the Schiff base. Above pH 6.5, the M-decay showed two phases. The fast M-decay was pH-independent and originated from the molecule having a protonated X-H, and the slow M-decay originated from the molecule having a deprotonated X, in which the proton came directly from the outside. The analysis yielded a value of 7.5 for the pKa of X-H. The proton uptake and release occurred during M-decay and O-decay, respectively.     

Remote Excitation Polarization-Dependent Surface Photochemical Reaction by Plasmonic Waveguide

For the first time, we report remote excitation polarization-dependent surface photochemical reaction by plasmonic waveguide. Remote excitation polarization-dependent surface-enhanced Raman scattering (SERS) spectra indicate a surface photochemical reaction that p-aminothiophenol is converted to p,p′-dimercaptoazobenzene (DMAB) induced by the plasmonic waveguide. Surface plasmon polaritons generated at the end of a silver nanowire can propagate efficiently along the nanowire, and be coupled by nanoparticles near the nanowire as a nanoantenna. Massive electromagnetic enhancement is generated in the nanogap between the nanowire and the nanoparticles. The remote excitation polarization-dependent SERS spectra can be obtained experimentally in the nanogaps; furthermore, the remote excitation polarization-dependent SERS spectra of DMAB reveal the occurrence of this surface catalytic reaction. Theoretical simulations using finite-difference time-domain methods strongly support our experimental results.     

Novel Proteorhodopsin variants from the Mediterranean and Red Seas

Proteorhodopsins, ubiquitous retinylidene photoactive proton pumps, were recently found in the widespread uncultured SAR86 bacterial group in oceanic surface waters. To survey proteorhodopsin diversity, new degenerate sets of proteorhodopsin primers were designed based on a genomic proteorhodopsin gene sequence originating from an Antarctic fosmid library. New proteorhodopsin variants were identified in Red Sea samples that were most similar to the original green-light absorbing proteorhodopsins found in Monterey Bay California. Unlike green-absorbing proteorhodopsins however, these new variants contained a glutamine residue at position 105, the same site recently shown to control spectral tuning in naturally occurring proteorhodopsins. Different proteorhodopsin variants were also found in the Mediterranean Sea. These proteorhodopsins formed new and distinctive proteorhodopsin groups. Phylogenetic analyses show that some of the new variants were very different from previously characterized proteorhodopsins, and formed the deepest branching groups found so far among marine proteorhodopsins. The existence of these varied proteorhodopsin sequences suggests that this class of proteins has undergone substantial evolution. These variants could represent functionally divergent paralogous genes, derived from the same or similar species, or orthologous proteorhodopsins that are distributed amongst divergent planktonic microbial taxa.     

Studies on a Missing Reaction in the Visual Cycle

DEVELOPMENT in the biochemistry of vision during the past twenty-five years can be summarized by equations (1) and (2) in Fig. 1, which envisage¹ that 11-cis-retinal combines with the visual protein opsin in a dark reaction to form the photolabile complex rhodopsin, λ_{max 497 nm}. When rhodopsin absorbs light it stimulates, through a process whose mechanism is not understood, the transmission of impulses, which are responsible for the visual sensation, although much is known about the biochemical changes accompanying the absorption of light by rhodopsin. These changes culminate in the formation of all-trans-retinal (λ_{max 385 nm}) and opsin (equation (2), Fig. 1), through a number of intermediates² and for the completion of the cycle one needs a molecular process which may regenerate 11-cis-retinal from all-trans-retinal (equation (3), Fig. 1).     

光化学反应诱导大鼠视网膜水肿模型

目的:通过静脉注射光敏剂Erythrosin B联合激光照射诱导大鼠视网膜水肿模型,并观察该模型中大鼠视网膜及血管形态改变.方法:大鼠尾静脉注射光敏剂Erythrosin B后,使用532nm Nd:YAG激光(1.95±0.05 mw)照射大鼠视网膜8分钟.分别于建模后第1、2、3、5、7及14天进行眼底照相、眼底荧光造影(FFA)及视网膜光学相干断层扫描(OCT)检查.并在OCT图像上测量大鼠视网膜厚度(RT).结果:眼底照相、FFA及OCT结果显示大鼠视网膜在激光照射后可立即出现血管损伤及视网膜厚度增加,但未发现视网膜血管栓塞.激光照射前RT为219±2 μm,激光照射后第1天RT即增加至283±6μm,并在第2天到达顶峰(302±7μm),之后开始下降,至第5天恢复到激光照射前水平(234±9 μm),到第14天时视网膜发生明显萎缩,RT较激光处理前减小(198±6μm).结论:这一通过光化学反应诱导的视网膜水肿模型具有可靠及可重复性,可被运用于视网膜水肿的病理学及动力学研究.     

Model Studies of the Iron-Catalysed Haber-Weiss Cycle and the Ascorbate-Driven Fenton Reaction

Complementary hydroxylation assays and stopped-flow e.s.r. techniques have been employed in the investigation of the effect of various iron chelators (of chemical, biological and clinical importance) on hydroxyl-radical generation via the Haber-Weiss cycle and the ascorbate-driven Fenton reaction.

Chelators have been identified which selectively promote or inhibit various reactions involved in hydroxyl-radical generation (for example, NTA and EDTA promote all the reactions of both the Haber-Weiss cycle and the ascorbate-driven Fenton reaction, whereas DTPA and phytate inhibit the recycling of iron in these reactions). The biological chelators succinate and citrate are shown to be relatively poor catalysts of the Haber-Weiss cycle, whereas they are found to be effective catalysts of ·OH generation in the ascorbate-driven Fenton reaction.

It is also suggested that continuous redox-cycling reactions between iron, oxygen and ascorbate may represent an important mechanism of cell death in biological systems.     

Model Studies of the Iron-Catalysed Haber-Weiss Cycle and the Ascorbate-Driven Fenton Reaction

Complementary hydroxylation assays and stopped-flow e.s.r. techniques have been employed in the investigation of the effect of various iron chelators (of chemical, biological and clinical importance) on hydroxyl-radical generation via the Haber-Weiss cycle and the ascorbate-driven Fenton reaction.

Chelators have been identified which selectively promote or inhibit various reactions involved in hydroxyl-radical generation (for example, NTA and EDTA promote all the reactions of both the Haber-Weiss cycle and the ascorbate-driven Fenton reaction, whereas DTPA and phytate inhibit the recycling of iron in these reactions). The biological chelators succinate and citrate are shown to be relatively poor catalysts of the Haber-Weiss cycle, whereas they are found to be effective catalysts of ·OH generation in the ascorbate-driven Fenton reaction.

It is also suggested that continuous redox-cycling reactions between iron, oxygen and ascorbate may represent an important mechanism of cell death in biological systems.     

Allostery Wiring Diagrams in the Transitions that Drive the GroEL Reaction Cycle

Determining the network of residues that transmit allosteric signals is crucial to understanding the function of biological nanomachines. During the course of a reaction cycle, biological machines in general, and Escherichia coli chaperonin GroEL in particular, undergo large-scale conformational changes in response to ligand binding. Normal mode analyses, based on structure-based coarse-grained models where each residue is represented by an α carbon atom, have been widely used to describe the motions encoded in the structures of proteins. Here, we propose a new C^α-side chain elastic network model of proteins that includes information about the physical identity of each residue and accurately accounts for the side-chain topology and packing within the structure. Using the C^α-side chain elastic network model and the structural perturbation method, which probes the response of a local perturbation at a given site at all other sites in the structure, we determine the network of key residues (allostery wiring diagram) responsible for the T → R and R″ → T transitions in GroEL. A number of residues, both within a subunit and at the interface of two adjacent subunits, are found to be at the origin of the positive cooperativity in the ATP-driven T → R transition. Of particular note are residues G244, R58, D83, E209, and K327. Of these, R38, D83, and K327 are highly conserved. G244 is located in the apical domain at the interface between two subunits; E209 and K327 are located in the apical domain, toward the center of a subunit; R58 and D83 are equatorial domain residues. The allostery wiring diagram shows that the network of residues are interspersed throughout the structure. Residues D83, V174, E191, and D359 play a critical role in the R″ → T transition, which implies that mutations of these residues would compromise the ATPase activity. D83 and E191 are also highly conserved; D359 is moderately conserved. The negative cooperativity between the rings in the R″ → T transition is orchestrated through several interface residues within a single ring, including N10, E434, D435, and E451. Signal from the trans ring that is transmitted across the interface between the equatorial domains is responsible for the R″ → T transition. The cochaperonin GroES plays a passive role in the R″ → T transition. Remarkably, the binding affinity of GroES for GroEL is allosterically linked to GroEL residues 350-365 that span helices K and L. The movements of helices K and L alter the polarity of the cavity throughout the GroEL functional cycle and undergo large-scale motions that are anticorrelated with the other apical domain residues. The allostery wiring diagrams for the T → R and R″ → T transitions of GroEL provide a microscopic foundation for the cooperativity (anticooperativity) within (between) the ring (rings). Using statistical coupling analysis, we extract evolutionarily linked clusters of residues in GroEL and GroES. We find that several substrate protein binding residues as well as sites related to ATPase activity belong to a single functional network in GroEL. For GroES, the mobile loop residues and GroES/GroES interface residues are linked.     

Inter-Ring Communication Is Dispensable in the Reaction Cycle of Group II Chaperonins

Chaperonins are ubiquitous molecular chaperones with the subunit molecular mass of 60 kDa. They exist as double-ring oligomers with central cavities. An ATP-dependent conformational change of the cavity induces the folding of an unfolded protein that is captured in the cavity. In the group I chaperonins, which are present in eubacteria and eukaryotic organelles, inter-ring communication takes important role for the reaction cycle. However, there has been limited study on the inter-ring communication in the group II chaperonins that exist in archaea and the eukaryotic cytosol. In this study, we have constructed the asymmetric ring complex of a group II chaperonin using circular permutated covalent mutants. Although one ring of the asymmetric ring complex lacks ATPase or ATP binding activity, the other wild-type ring undergoes an ATP-dependent conformational change and maintains protein-folding activity. The results clearly demonstrate that inter-ring communication is dispensable in the reaction cycle of group II chaperonins.     

紫细菌光合反应中心中细菌脱镁叶绿素被置换后的光化学活性

采用新型表面活性剂ＬＤＡＯ,结合ＤＥＡＥ纤维素层析法,我们提纯了紫细菌Ｒｈｏｄｏｂａｃｔｅｒｓｐｈａｅｒｏｉｄｅｓ６０１的光合反应中心。在一定温度和丙酮的协同作用下,外加的植物脱镁叶绿素ａ可取代反应中心细菌脱镁叶绿素,形成含有脱镁叶绿素ａ的紫细菌光合反应中心（ＰｈｅａＲＣ）。当协同作用１５ｍｉｎ和６０ｍｉｎ时,反应中心中的细菌脱镁叶绿素分别被替代了５０％和７１％。在ＰｈｅａＲＣ中,细菌脱镁叶绿素的ＱＸ（５３７ｎｍ）和ＱＹ（７５８ｎｍ）特征峰显著下降,而出现高等植物脱镁叶绿素的ＱＸ（５０９／５４２ｎｍ）和ＱＹ（６７４ｎｍ）特征峰。排除温度和丙酮的影响,替代时间为１５ｍｉｎ或６０ｍｉｎ的ＰｈｅａＲＣ的光化学活性分别为对照的７８％或７１％。     

变形菌视紫红质十年的回顾与展望

变形菌视紫红质(proteorhodopsin,PR)是一类吸光色素膜蛋白,它是由7个跨膜ɑ-螺旋(A～G)组成的视蛋白(opsins)与色素视黄醛(retinal)通过共价键结合而形成。PR广泛存在于海洋和淡水水域的微生物中,是一种质子泵型的视紫红质,可在光的驱动下将质子从细胞质泵到胞质间隙,从而在细胞内外产生质子梯度,形成的化学势能被用于合成ATP、物质的跨膜运输以及驱动鞭毛运动等方面。据估算含PR的细菌在海洋水域中约占总细菌的13%,而每个含PR的细胞中的PR分子数约为2.5×104个。对PR功能的研究表明PR具有增强其宿主菌抵抗外界不良环境的能力;而对于PR三维结构的研究对PR的作用机理及其功能的研究将有更进一步的推动作用。     

Lipid Bilayer Composition Can Influence the Orientation of Proteorhodopsin in Artificial Membranes

Artificial membrane systems allow researchers to study the structure and function of membrane proteins in a matrix that approximates their natural environment and to integrate these proteins in ex vivo devices such as electronic biosensors, thin-film protein arrays, or biofuel cells. Given that most membrane proteins have vectorial functions, both functional studies and applications require effective control over protein orientation within a lipid bilayer. In this work, we explored the role of the bilayer surface charge in determining transmembrane protein orientation and functionality during formation of proteoliposomes. We reconstituted a model vectorial ion pump, proteorhodopsin, in liposomes of opposite charges and varying charge densities and determined the resultant protein orientation. Antibody-binding assay and proteolysis of proteoliposomes showed physical evidence of preferential orientation, and functional assays verified the vectorial nature of ion transport in this system. Our results indicate that the manipulation of lipid composition can indeed control orientation of an asymmetrically charged membrane protein, proteorhodopsin, in liposomes.     

Lipid Bilayer Composition Can Influence the Orientation of Proteorhodopsin in Artificial Membranes

Artificial membrane systems allow researchers to study the structure and function of membrane proteins in a matrix that approximates their natural environment and to integrate these proteins in ex vivo devices such as electronic biosensors, thin-film protein arrays, or biofuel cells. Given that most membrane proteins have vectorial functions, both functional studies and applications require effective control over protein orientation within a lipid bilayer. In this work, we explored the role of the bilayer surface charge in determining transmembrane protein orientation and functionality during formation of proteoliposomes. We reconstituted a model vectorial ion pump, proteorhodopsin, in liposomes of opposite charges and varying charge densities and determined the resultant protein orientation. Antibody-binding assay and proteolysis of proteoliposomes showed physical evidence of preferential orientation, and functional assays verified the vectorial nature of ion transport in this system. Our results indicate that the manipulation of lipid composition can indeed control orientation of an asymmetrically charged membrane protein, proteorhodopsin, in liposomes.     

Photochemical Reaction Dynamics of the Primary Event of Vision Studied by Means of a Hybrid Molecular Simulation

The photoisomerization reaction dynamics of a retinal chromophore in the visual receptor rhodopsin was investigated by means of hybrid quantum mechanical/molecular mechanical (QM/MM) molecular dynamics (MD) simulations. The photoisomerization reaction of retinal constitutes the primary step of vision and is known as one of the fastest reactions in nature. To elucidate the molecular mechanism of the high efficiency of the reaction, we carried out hybrid ab initio QM/MM MD simulations of the complete reaction process from the vertically excited state to the photoproduct via electronic transition in the entire chromophore-protein complex. An ensemble of reaction trajectories reveal that the excited-state dynamics is dynamically homogeneous and synchronous even in the presence of thermal fluctuation of the protein, giving rise to the very fast formation of the photoproduct. The synchronous nature of the reaction dynamics in rhodopsin is found to originate from weak perturbation of the protein surroundings and from dynamic regulation of volume-conserving motions of the chromophore. The simulations also provide a detailed view of time-dependent modulations of hydrogen-out-of-plane vibrations during the reaction process, and identify molecular motions underlying the experimentally observed dynamic spectral modulations.