A [Ni(cyclam)]2+-functionalised poly(3,4-ethylenedioxythiophene): Synthesis,electropolymerisation and characterisation of the polymer by cyclic voltammetry and in situ reflectance FTIR spectroscopy

A 1,4,8,11-tetraazacyclotetradecane (cyclam) derivative of 3,4-ethylenedioxythiophene (EDOT) has been synthesised. Its square planar Ni(II) tetrafluoroborate complex has been electrochemically polymerised, with EDOT itself, to give a highly stable conducting polymer with covalently attached [Ni(cyclam)]²⁺ moieties. The cyclic voltammogram shows redox behaviour typical of a functionalised PEDOT, with the reversible Ni(II)–Ni(III) process of the [Ni(cyclam)]²⁺ complex superimposed. Reflectance in situ FTIR spectroscopy (RIFTIRS) shows that the presence of the metal complex has a profound influence upon the behaviour of the electronic band of the oxidised form of the polymer, and of the vibrational signature due to the charge carriers.     

Biomimetic synthesis of a water soluble conducting molecular complex of polyaniline and lignosulfonate

A new biomimetic route for the synthesis of a conducting molecular complex of polyaniline (Pani) and a natural polyelectrolyte, lignosulfonate (LGS) is presented. A poly(ethylene glycol) modified hematin (PEG-hematin) was used to catalyze the polymerization of aniline in the presence of LGS to form a Pani/LGS complex. UV-vis, FTIR, conductivity and TGA studies for the LGS-polyaniline complex indicate the presence of a thermally stable and electrically conductive form of polyaniline. Also the presence of LGS in this complex, an inexpensive byproduct from pulp processing, provides a unique combination of properties such as electronic conductivity, processability and biodegradability. The use of this conductive complex for corrosion protection is also proposed.     

蔗糖磷酸合酶功能、结构与催化机制的研究进展

蔗糖是自然界中广泛存在的一种天然产物.在植物等生命体中,蔗糖磷酸合酶(Sucrose phosphate synthase,SPS)是蔗糖合成的限速酶.SPS催化合成蔗糖-6-磷酸;蔗糖磷酸酶(Sucrose Phosphatase,SPP)进一步把蔗糖-6-磷酸上的磷酸根水解下来而形成蔗糖.近几十年来关于SPS的研究...     

The location of DNA in complexes of recA protein with double-stranded DNA. A neutron scattering study

Purified recA protein is found as rodlike homopolymers, and it forms filamentous complexes with double-stranded DNA that are stable in the presence of ATP gamma S, a nonhydrolyzable analogue of ATP. The structure of these filaments has been described in some detail by electron microscopy. Here we confirm the mass per length of 6.5 recA/100 A in solution by small-angle neutron scattering and extend the analysis to homopolymers of recA protein, finding a mass per length of about 7 recA/100 A and a radial mass distribution (cross-sectional radius of gyration) significantly different for the two filaments. The models proposed so far for the structure of the complex have placed the DNA in the center of the filament. Here we verify this assumption using small-angle neutron scattering to locate the DNA in the complexes, exploiting the contrast variation method in D2O/H2O mixtures. Model calculations show that the natural contrast difference between DNA and protein is not sufficient to locate the DNA (which accounts for only 4.7% of the mass in the complex). When deuterated DNA is used, the contrast difference is enhanced, and model calculations and experiment then converge, indicating that the DNA is indeed near the axis of the complex.     

Synthesis,copolymerization and peptide-modification of carboxylic acid-functionalized 3,4-ethylenedioxythiophene (EDOTacid) for neural electrode interfaces

Background

Conjugated polymers have been developed as effective materials for interfacing prosthetic device electrodes with neural tissue. Recent focus has been on the development of conjugated polymers that contain biological components in order to improve the tissue response upon implantation of these electrodes.

Methods

Carboxylic acid-functionalized 3,4-ethylenedioxythiophene (EDOTacid) monomer was synthesized in order to covalently bind peptides to the surface of conjugated polymer films. EDOTacid was copolymerized with EDOT monomer to form stable, electrically conductive copolymer films referred to as PEDOT-PEDOTacid. The peptide GGGGRGDS was bound to PEDOT-PEDOTacid to create peptide functionalized PEDOT films.

Results

The PEDOT-PEDOTacid-peptide films increased the adhesion of primary rat motor neurons between 3 and 9 times higher than controls, thus demonstrating that the peptide maintained its biological activity.

Conclusions

The EDOT-acid monomer can be used to create functionalized PEDOT-PEDOTacid copolymer films that can have controlled bioactivity.

General Significance

PEDOT-PEDOTacid-peptide films have the potential to control the behavior of neurons and vastly improve the performance of implanted electrodes. This article is part of a Special Issue entitled Organic Bioelectronics—Novel Applications in Biomedicine.     

Growth of thin processable films of poly(pyrrole) using phosphomolybdate clusters

Facile control over the chemical polymerization of pyrrole from solutions of pyrrole-phosphomolybdic acid has allowed processibility of conducting poly(pyrrole). This approach has allowed thin conducting films of poly(pyrrole) to be deposited in a single step, and in contrast to electrochemical polymerization methods, is not limited to electrically conducting substrates. For example, 40–100 nm thick poly(pyrrole) films with conducivities in the range of 15 to 30 S cm⁻¹ have been spin coated onto insulating glass substrates. Insight into the mechanism of polymerization has been obtained with cyclic voltammetry and UV-Vis spectroscopy.     

Topological mapping of the active sites of cytochromes P4502B1 and P4502B2 by in situ rearrangement of aryl-iron complexes.

Cytochrome P4502B1 reacts with phenylhydrazine or phenyldiazene to give an iron-phenyl complex that oxidatively rearranges in situ to the two N-phenylprotoporphyrin IX regioisomers with the phenyl group on pyrrole rings A (NA) and D (ND) [Swanson, B. A., Dutton, D. R., Lunetta, J. M., Yang, C. S., & Ortiz de Montellano, P. R. (1991) J. Biol. Chem. 266, 19258-19264]. The conclusion that the active site of cytochrome P4502B1 is open above pyrrole rings A and D but not B and C is extended here by studies with larger arylhydrazines. The N-arylprotoporphyrin IX standards required for product identification were obtained by reaction of the arylhydrazines with equine myoglobin. Cytochrome P4502B1 aryl-iron complex formation followed by oxidative shift of the aryl group produces the following N-aryl-protoporphyrin IX NA:ND regioisomer ratios: phenylhydrazine (39:61), 3,5-dimethylphenylhydrazine (29:71), 4-tert-butylhydrazine (25:75), 2-naphthylhydrazine (less than 2:greater than 98), and 4-(phenyl)phenylhydrazine (87:13). Electron-withdrawing substituents (as in 3,5-dichlorophenyl) prevent the aryl group shift. The increase in the proportion of the ND regioisomer with increasing bulk of the aryl group suggests that the region over pyrrole ring A is more sterically encumbered than that over pyrrole ring D. The regiospecificity is reversed, however, with 4-(phenyl)phenylhydrazine, which primarily gives the NA regioisomer. This reversal suggests that the active site has a sloping roof that is higher over pyrrole ring A than pyrrole ring D and that provides a larger steric barrier to the shift of tall aryl moieties than the barrier over pyrrole ring A.(ABSTRACT TRUNCATED AT 250 WORDS)     

Characterization of a rice sucrose-phosphate synthase-encoding gene

A rice genomic clone (spsl) coding for sucrose phosphate synthase (SPS) was isolated and sequenced. Rice spsl contains 13 exons and 12 introns, an unusually long 366-bp leader region with a highly organized primary structure and a promoter region with no obvious homology with eukaryotic promoter consensus sequences. Southern blot analysis showed that SPS is encoded by a single-copy gene in the rice genome. Comparison of the rice, maize, potato and spinach SPS deduced amino acid (aa) sequences showed that these enzymes have a well conserved region comprising their first 700 aa, and a variable C-terminal region. Analysis of rice spsl expression showed that mRNA levels change during leaf development. SPS activity and mRNA were undetectable in roots.     

Poly(dG).poly(dC) at neutral and alkaline pH: the formation of triple stranded poly(dG).poly(dG).poly(dC).

Alkaline titrations of different samples of poly(dG).poly(dC) and of the constituent homopolymers poly(dG) and poly(dC) have been performed in 0.15 M NaCl and their CD spectra followed. Sample I contained a slight excess of poly(dC) (52% C: 48% G) and showed a single reversible transition (pK = 11.9) due to the dissociation of double stranded poly(dG).poly(dC). Sample II, containing an excess of poly(dG) (43% C: 57% G), showed two transitions (pK1 = 11.4, PK2 = 11.9) the first one being only partially reversible. Examination of the CD spectra along the alkaline titrations indicated the presence of another hydrogen-bonded complex of higher G content. Mixing curves performed at pH 8 have confirmed the presence of a 2G: 1C complex, besides the double stranded complex. It can be formed in amounts up to 30% by mixing the two homopolymers, alkali treatment and heating. The CD spectra of the two complexes have been computed from the CD data of the mixing curves. This permitted the determination of the concentrations of both complexes and homopolymers in all samples. The ratio of triple to double stranded complex is not only dependent on the G/C ratio of the sample, but also a function of the previous physico-chemical conditions. These results explain the variability of many properties of different poly(dG).poly(dC) samples observed by other workers.     

A new copper(II) complex of unsymmetrical tetradentate ligand generated in situ: synthesis and molecular structure

A new copper(II) complex with tetradentate unsymmetrical ligand was prepared by one-pot condensation of methyl-2-pyrrole carboxylate, diethylenetriamine and copper(II) sulfate. The complex was characterized by elemental analysis, electronic and IR spectral, as well as X-ray crystal structure determination. The X-ray structure of the molecule reveals the copper(II) center is in a square planar environment through coordination by two nitrogen atoms of the amine, one amide nitrogen atom and one nitrogen atom of the pyrrole moieties, respectively. The copper(II) complex is neutral due to deprotonation of the amide and pyrrole groups.     

Physical and Kinetic Evidence for an Association between Sucrose-Phosphate Synthase and Sucrose-Phosphate Phosphatase

The possible formation of a multienzyme complex between sucrose (Suc)-phosphate synthase (SPS) and Suc-phosphate phosphatase (SPP) was examined by measuring the rates of Suc-6-phosphate (Suc-6-P) synthesis and hydrolysis in mixing experiments with partially purified enzymes from spinach (Spinacia oleracea) and rice (Oryza sativa) leaves. The addition of SPP to SPS stimulated the rate of Suc-6-P synthesis. SPS inhibited the hydrolysis of exogenous Suc-6-P by SPP when added in the absence of its substrate (i.e. UDP-glucose) but stimulated SPP activity when the SPS substrates were present and used to generate Suc-6-P directly in the reaction. Results from isotope-dilution experiments suggest that Suc-6-P was channeled between SPS and SPP. A portion of the SPS activity comigrated with SPP during native polyacrylamide gel electrophoresis, providing physical evidence for an enzyme-enzyme interaction. Taken together, these results strongly suggest that SPS and SPP associate to form a multienzyme complex.     

Unusual regulatory properties of sucrose-phosphate synthase purified from soybean (Glycine max) leaves

Sucrose-phosphate (SPS) from source leaves of soybean ( Glycine max (L.) Merr. cv. Ransom II) was purified 74-fold to a final specific activity of 1.8 U (mg protein)¹. The partially purified preparation was free from phosphoglucoseisomerase (EC 5.3.1.9), pyrophosphatase (EC 3.6.1.1), phosphoenolpyruvate-phosphatase (EC 3.1.3.-), phosphofructokinase (EC 2.7.1.11), and uridine diphosphatase (EC 3.6.1.6), and was used for characterization of the kinetic and regulatory properties of the enzyme. The enzyme showed hyperbolic saturation kinetics for both fructose-6-phosphate (K_m=0.57 m M ) and UDPGlucose (UDPG) (K_m=4.8 m M ). The activity of SPS was inhibited by the product UDP. In vitro this inhibition could be partially overcome by the presence of Mg²⁺. Inorganic orthophosphate was only slightly inhibitory (35% inhibition at 25 m M phosphate). Glucose-6-phosphate (up to 20 m M ) had no effect on activity, and did not show any significant interaction with phosphate inhibition. A range of potential effectors was tested and had no effect on SPS activity: Glucose-1-phosphate, fructose-1, 6-bisphosphate, α-glycero-phosphate, dihydroxyacetone-phosphate, 3-phosphoglyceric acid, (all at 5 m M ), sucrose at 100 m M and pyrophosphate at 0.1 m M . The apparent lack of allosteric regulation of soybean SPS makes this enzyme markedly different from SPS previously characterized from spinach and maize.     

If the antibody fails--a mass western approach

SUMMARY: Sucrose-phosphate synthase (SPS) has attracted the interest of plant scientists for decades. It is the key enzyme in sucrose metabolism and is under investigation in various plant species, e.g. spinach, tobacco, poplar, resurrection plants, maize, rice, kiwi and Arabidopsis thaliana. In A. thaliana, there are four distinct SPS isoforms. Their expression is thought to depend on environmental conditions and plant tissue. However, these data were derived from mRNA expression levels only. No data on SPS protein identification from crude extracts have been available until now. An antibody approach failed to distinguish the four isoforms. Therefore, we developed a method for SPS quantification and isoform-specific identification in A. thaliana complex protein samples. Samples were separated on SDS-PAGE, digested and directly applied to liquid chromatography/triple-stage quadrupole mass spectrometry (LC/TSQ-MS). In this approach, known as mass Western, samples were analysed in multi-reaction monitoring (MRM) mode, so that all four SPS isoforms could be measured in one experiment. In addition to the relative quantification, stable isotope-labelled internal peptide standards allowed absolute quantification of SPS proteins. Protein extracts from various plant tissues, samples harvested during the day or the night, and cold-stressed plants were analysed. The stress-specific SPS5a isoform showed increased concentrations in cold-stressed leaf material.     

Formation of and coalescence from the inclusion complex of a biodegradable block copolymer and alpha-cyclodextrin. 2: A novel way to regulate the biodegradation behavior of biodegradable block copolymers

A biodegradable block copolymer (PCL-b-PLLA, M(n) = 1.72 x 10(4), M(w)/M(n) = 1.37) of poly(epsilon-caprolactone) (PCL) and poly(L-lactide) (PLLA) with very low crystallinity was obtained by forming the inclusion complex between alpha-cyclodextrin molecules and PCL-b-PLLA followed by coalescence of the guest polymer chains. Films of the as-synthesized and coalesced copolymer samples, PCL and PLLA homopolymers of approximately the same chain lengths as the corresponding blocks of PCL-b-PLLA, and a physical blend of PCL/PLLA homopolymers with the same molar composition as PCL-b-PLLA were prepared by melt-compression molding between Teflon plates. Subsequently, the in vitro biodegradation behavior of these films was studied in phosphate buffer solution containing lipase from Rhizopus arrhizus, by means of ultraviolet spectra, attenuated total reflectance FTIR spectra, differential scanning calorimetry, wide-angle X-ray diffraction measurements, and weight loss analysis. PCL segments were found to degrade much faster than PLLA segments, both in the pure state and in copolymer or blend samples. Consistent with our expectation, suppression of the phase separation, as well as a decrease of crystallinity, in the coalesced copolymer sample led to a much faster enzymatic degradation than that of either as-synthesized copolymer or the PCL/PLLA physical blend sample, especially during the early stages of biodegradation. Thus the biodegradation behavior of biodegradable block copolymers, which is of decisive importance in drug delivery and controlled release systems, may be regulated by the novel and convenient means recently reported by us.(1)     

Theoretical investigation on the selective detection of SO2 molecule by AlN nanosheets

A density functional theory investigation of adsorption of monomer, dimer and trimer forms of pyrrole on the outer surface of zigzag (7,0) single-walled carbon nanotube (SWCNT) has been reported. Geometries of the complexes were optimized using the M06-2X functional and the 6-31G(d,p) basis set. Moreover, 6-311G(d,p), cc-pVDZ and cc-pVTZ basis sets were used for the adsorption energy calculation and such energies were corrected for the basis set superposition error. Vertical ionization potential and electron affinity of the investigated system were also computed. The interaction of polypyrrole on the SWCNT surface is characterized by the stacking interaction. Adsorption (binding) energy of pyrrole on the SWCNT surface is weak, but such energy increases with the number of monomer units in the pyrrole oligomer. In the SWCNT-pyrrole complexes, the oxidation and reduction processes will take place only at the SWCNT. The influence of larger unit on the electronic properties of the complex has been detailed.     

Production of Slime Polysaccharides by Shigella dysenteriae Type 1

Electron microscopy of ruthenium red-stained ultrathin section of strains of Shigella dysenteriae type 1 grown in the Casamino Acids-yeast extract broth medium showed the presence of an extracellular slime layer. The slime appeared as a dense sheath covering bacteria. The presence of slime promoted hemagglutinating activity of the bacteria. The slime polysaccharide (SPS) isolated from the cell-free culture supernatant or the bacterial surface was less than 162,000 daltons in size and immunochemically similar. The SPS showed cross-reaction with lipopolysaccharide (LPS) antigen in immunological tests; however, it also appeared to be different from LPS since it did not contain 2-keto-3-deoxyoctonate, a core sugar of LPS. A different pattern of separation from LPS was also observed by silver staining of SDS-polyacrylamide gels. From these data it appeared that either LPS and SPS are contaminated with each other or that SPS is the polysaccharide portion of LPS.     

Phosphorylation of synthetic peptides by a CDPK and plant SNF1-related protein kinase. Influence of proline and basic amino acid residues at selected positions.

Spinach (Spinacia oleracea L.) leaf sucrose-phosphate synthase (SPS) can be inactivated by phosphorylation of Ser-158 by calmodulin-like domain protein kinases (CDPKs) or SNF1-related protein kinases (SnRK1) in vitro. While the phosphorylation site sequence is relatively conserved, most of the deduced sequences of SPS from dicot species surrounding the Ser-158 regulatory phosphorylation site contain a Pro residue at P-4 (where P is the phosphorylated Ser); spinach is the exception and contains an Arg at P-4. We show that a Pro at P-4 selectively inhibits phosphorylation of the peptide by a CDPK relative to a SnRK1. The presence of a Pro at P-4, by allowing a tight turn in the peptide substrate, may interfere with proper binding of residues at P-5 and beyond. Both kinases had greater activity with peptides having basic residues at P-6 and P+5 (in addition to the known requirement for an Arg at P-3/P-4), and when the residue at P-6 was a His, the pH optimum for phosphorylation of the peptide was acid shifted. The results are used to predict proteins that may be selectively phosphorylated by SnRK1s (as opposed to CDPKs), such as SPS in dicot species, or may be phosphorylated in a pH-dependent manner.     

The 59-kDa polypeptide constituent of 8–10-nm cytoplasmic filaments in Neurospora crassa is a pyruvate decarboxylase

The fungus Neurospora crassa harbors large amounts of cytoplasmic filaments which are homopolymers of a 59-kDa polypeptide (P59Nc). We have used molecular cloning, sequencing and enzyme activity measurement strategies to demonstrate that these filaments are made of pyruvate decarboxylase (PDC, EC 4.1.1.1), which is the key enzyme in the glycolytic-fermentative pathway of ethanol production in fungi, and in certain plants and bacteria. Immunofluorescence analyses of 8–10-nm filaments, as well as quantitative Northern blot studies of P59Nc mRNA and measurements of PDC activity, showed that the presence and abundance of PDC filaments depends on the metabolic growth conditions of the cells. These findings may be of relevance to the biology of ethanol production by fungi, and may shed light on the nature and variable presence of filament bundles described in fungal cells.