Kouji Iida

Photocurrent and electronic activities of oriented-His-tagged photosynthetic light-harvesting/reaction center core complexes assembled onto a gold electrode.

A polyhistidine (His) tag was fused to the C- or N-terminus of the light-harvesting (LH1)-α chain of the photosynthetic antenna core complex (LH1-RC) from Rhodobacter sphaeroides to allow immobilization of the complex on a solid substrate with defined orientation. His-tagged LH1-RCs were adsorbed onto a gold electrode modified with Ni-NTA. The LH1-RC with the C-terminal His-tag (C-His LH1-RC) on the modified electrode produced a photovoltaic response upon illumination. Electron transfer is unidirectional within the RC and starts when the bacteriochlorophyll a dimer in the RC is activated by light absorbed by LH1. The LH1-RC with the N-terminal His-tag (N-His LH1-RC) produced very little or no photocurrent upon illumination at any wavelength. The conductivity of the His-tagged LH1-RC was measured with point-contact current imaging atomic force microscopy, indicating that 60% of the C-His LH1-RC are correctly oriented (N-His 63%). The oriented C-His LH1-RC or N-His LH1-RC showed semiconductive behavior, that is, had the opposite orientation. These results indicate that the His-tag successfully controlled the orientation of the RC on the solid substrate, and that the RC produced photocurrent depending upon the orientation on the electrode.

Liposomal polyamine-dialkyl phosphate conjugates as effective gene carriers: chemical structure, morphology, and gene transfer activity.

Synthetic cationic lipids are promising transfection agents for gene therapy. We report here that polyamine conjugates of dialkyl phosphates, combined with natural lipids and assembled in the form of liposomes (polycationic liposome: PCL), possess high transfection activity in the COS-1 cell line. Furthermore, we describe the functional morphology of the PCL/DNA complexes as revealed by atomic force microscopy (AFM). The conjugates were synthesized from dialkyl phosphates (with alkyl chain lengths of 12, 14, or 16 carbons) by reaction with the polyamine molecules, spermidine, spermine, or polyethylenimine (PEI(1800)). [Dewa, T., et al. Bioconjugate Chem. 2004, 15, 824]. The PCL composed of the spermidine and C16 conjugate combined with phospholipid and cholesterol (conjugate/phospholipid/cholesterol = 1/1/1 as a molar ratio) exhibited 3.6 times higher activity than that of a popular commercial product. Systematic tests revealed clear correlations of the transgene activity with physical properties of the polyamine, in particular, that longer alkyl chains and the lower molecular weight polyamines (spermidine, spermine) favor high efficacy at the higher nitrogen/phosphate ratio = 24 (N/P, stoichiometric ratio of nitrogen in the conjugate to phosphate in DNA). The low molecular weight polyamine-based PCLs, which formed 150-400 nm particles with plasmid DNA (lipoplexes), exhibited approximately 3-fold higher gene transfer activity than micellar aggregates (lacking phospholipid and cholesterol) of the corresponding conjugate. In contrast, the PEI-based PCL formed large aggregates (approximately 1 microm), that, like the micellar aggregate form, had low activity. Activity of the low molecular weight polyamine-based PCLs increased linearly with the N/P of the lipoplex up to N/P = 24. Formation of lipoplexes was examined by agarose gel electrophoresis, dynamic light scattering (DLS), and AFM. At the lower N/P = 5, large aggregates of complex (approximately 1 microm), in which DNA molecules were loosely packed, were observed. At higher N/P, lipoplexes were converted into smaller particles (150-400 nm) having a lamellar structure, in which DNA molecules were tightly packed. Such morphological features of the lipoplex correlate with the dependence of transfection on the N/P in that the lamellar structures gave superior transfection. AFM also indicated that the lipoplexes disassembled significantly, releasing DNA, when the lipoplexes were exposed to acidic conditions (pH 4). The significance for transfection activity of the metamorphosis of bilayer lipoplexes is discussed relative to that of the less active micellar aggregate form, which is unresponsive to pH change.

Electron Transfer Mediated by Photosynthetic Reaction Center Proteins between Two Chemical-Modified Metal Electrodes

ABSTRACT Photosynthetic reaction center (RC) proteins in photosynthetic bacteria are very interesting from the viewpoint of natural supramolecular electric devices. One of key factors for the enhancement in currents of RC protein devices is the achievement of low electron-injection barrier between proteins and metal electrodes. In this study, the electron transfer mediated by RC proteins between two gold electrodes modified with various self assemble monolayer (SAM) materials, was measured by use of conductive atomic force microscopy (CAFM). We found that, when both the Au(111) metal substrate and the gold-coated cantilever were modified with 2-mercaptopyridine, CAFM currents were especially enhanced.

Molecular assembly of covalently-linked mesoporphyrin dimers with light-harvesting polypeptides

Abstract Light-harvesting (LH)-α and -β polypeptides separately isolated from photosynthetic bacteria, R. rubrum , organized mesoporphyrin dimers ( Scheme 1 ) in the n -octyl-β- d -glucopyranoside (OG) micelle, depending upon the porphyrin structure and temperature. An efficient energy transfer from Zn porphyrin to Ni porphyrin in 7 due to the presence of the LH polypeptides was observed.

Synthesis and characterization of soluble polymer ligands possessing sulfide side chains

Abstract Soluble polymer ligands possessing aryl monosulfide side chains [poly(4-vinylphenylmethyl sulfide) (PVPMS)], alkyl monosulfide side chains [poly(4-vinylbenzylmethyl sulfide) (PVBMS)] and branched tetrasulfide side chains [poly(7-(4′-vinylbenzyloxy)-2,5,9,12-tetrathiatridecane) (PTeSSD)] were prepared by radical polymerization of the corresponding monomers. These polymer ligands had specific binding abilities for soft acids only, such as Ag(I) and Hg(II) ions. The tetrasulfide-type polymer ligand, PTeSSD, bound the metal ion only by the tetrasulfide side chain itself. The monosulfide-type polymer ligands, PVPMS and PVBMS, bound the metal ion by co-operative interaction between their monosulfide side chains. The resulting polymer-Ag(I) complexes were soluble in organic solvents. Interestingly, a polyester cloth coated with the polymer-Ag(I) complex exhibited significant antibacterial activity against Staphylococcus aureus .

Two-dimensional patterning of bacterial light-harvesting 2 complexes on lipid-modified gold surface

In a photosynthetic membrane, nano-scale patterns of light-harvesting (LH) pigment-protein complexes play an essential role in capturing photons and ensuring efficient excitation energy transfer. LH complexes 1 and 2 have drawn attention as building blocks of a nano-scale photoelectric device. For obtaining a device with efficiency comparable to that of the natural photosynthesis, a method has to be established for forming a two-dimensional assembly of LH complexes around a metal electrode. In this study, LH2 complexes isolated from Rb. sphaeroides were immobilized on a patterned gold surface. Quenching of photo-excitation energy by gold was prevented through the placement of a self-assembled phospholipid monolayer between the LH2 complexes and the gold surface.

Construction of Langmuir–Blodgett films from light-harvesting complex I isolated from photosynthetic bacteria

Abstract Langmuir–Blodgett (LB) films of the light-harvesting (LH) polypeptides/bacteriochlorophyll a (BChl a) complex isolated from photosynthetic bacteria, Rhodospirillum rubrum, were constructed. The monolayers of the core LH /BChl a complex were prepared from octyl β-glycoside suspensions at an air/water interface and then the monolayer was transferred to glass slides using LB technique multilayer films. UV-vis absorption spectra of the LB layer films absorbing 870 nm at the Qy absorption of BChl a indicated that the LH/BChl a complex in the multilayer films was more stable than the subunit complex absorbing 820 nm at the Qy absorption. Analysis of Fourier transform infrared reflection absorption spectroscopy (FTIR-RAS) spectra showed that the tilt angle of the transmembrane axis of the LH/BChl a complex in the multilayer films was 40° to the normal.

Assembly of Bacterial Light-Harvesting Complexes on Solid Substrates

Light-harvesting (LH) and reaction center (RC) complexes were adsorbed onto mica and gold substrates, in order to assemble an artificial antenna complex on a solid support, with the eventual aim of developing useful nanodevices. The normal near-infra-red (IR) absorption spectra of photosynthetic complexes were retained indicating stable assembly on these solid substrates. Atomic Force Microscopy (AFM) of reassociated LH1 complexes, under air, showed the expected ring-like structure.