Interaction Between HMG Proteins (1+2) and the Negative Regulatory Region 1(NCR1) in the 5''''-flanking Sequence of the Human β-globin Gene

The pattern of high mobility group proteins 1 and 2 (HMG1,2) interaction with the 5'-flanking sequence of the human β-globin gene has been analyzed by scanning tunnelling microscopy (STM). A 200 bp negative regulatory region in the 5'-flanking sequence of the human β-globin gene can be folded by HMG proteins 1 and 2 into a circular structure (diameter 70±6) with a linear tail which seems to be a left-handed double helix structure.     

CLONING, EXPRESSION AND CHARACTERIZATION OF Pro3 GENE OF YEAST Saccharomyces cerevisiae

The proline biosynthetic pathway and Pro genes in Saccharomyces cerevisiae have just begun to be studied recently. In our laboratory, Pro2 gene of S. cerevisiae had been cloned in yeast. As described in this paper, yeast Pro3 gene was also cloned, which can complement yeast Pro3 mutants, and be expressed efficiently in E. coli. The high activities of this gene product, L-pyrroline-5-carboxylate (P5C) reductase, can be detected in both organisms. The activity of the Pro3 gene product in multiple copy plasmids is not higher than that of single copy genes in chromosomes in both yeast and E. coll. The preliminary characterization of the gene is also reported.     

Fluorescent Switch Behavior of 1, 2-Bis（4-pyridyl） Ethylene Controlled by pH in Aqueous Solution

With the decrease of pH value from 8.45 to -1.0, the UV-Vis absorption and fluorescent spectra of 1,2-bis（4-pyridyl） ethylene（BPE） took on the same changing trend at four different successive pH stages： 8.45--7.20, 7.20--5.62, 5.62--2.60, and 2.60--1.0, namely, no change, decrease, increase, and decrease again. Among these, in a range of 7.20--5.62, the fluorescence wavelength blueshifted from 418 to 359 nm, but the UV-Vis absorption wavelength, in contrast, redshifted from 285 to 298 nm. The fluorescence intensity of BPE had a drop even to quench upon a decline in the pH value from 2.60 to -1.0 probably owing to its cation-re interaction to reduce the π electron cloud density of BPE. Two dissociation constants, pKa1（4.30±0.01） and PKa2（5.65±0.04）, were obtained based on fluorescence data. The changes of fluorescence spectra indicate that BPE has ＂oft-on-off＂ switch behavior. The fluorescent spectra of BPE were nearly independent on the presence of α- and β-cyclodextrins.     

Structure Analysis of Streptococcal Protein G Fc Binding Domain

The gene fragment (191 bp) encoding protein G IgG Fc binding domain was isolated by PCR from group G streptococcus (CMCC32138), and a clone containing this gene fragment was found to give fine reactivity to human IgG when expressed in Escherichia coli. The complete nucleotide sequence of the gene fragment was determined. One base pair differs from previously reported protein Gnucleotide sequences, and resultsin an amino acid change (Ala-Thr), but this variation makes no difference in binding to the IgG Fc part by ELISA.The secondary structure of the protein G IgG Fc binding domain has been estimated by circular dichroism and assigned by computer algorithm.It shows a typical α-helix region in this domain.By breaking this α-helix region with recombinant DNA techniques, a 44 peptide, which contained the N-terminal 27 amino acid residues of this domain, was expressed in E. coli and showed no reactivity to IgG.The hydropathicity of this domain was also analyzed and compared with that of protein A relevant     

RECOGNITION SEQUENCE SPECIFICITY OF SIGNAL PEPTIDASE I AND THE ROLE OF SIGNAL PEPTIDE IN SECRETION OF PROTEIN IN Bacillus subtilis

By recombinant DNA technology, the N-terminal of the β-protein encoding region of plasmid pUBHO is fused with the structure gene of α-amylase from Bacillus licheniformis. This gene fusion is called βAmy. It is able to transcribe and translate in phase. Protein fusion can be secreted into the medium mediated by β-signal peptide. The efficiency of secretion is about 10% of the synthesized pre-α-amylase. By comparing the secretion capacities and analysing the restriction sites on β-Amy genes and the molecular weights of the mature α-amylase secreted by B. subtilis harbouring different plasmids, it is indicated in vivo that the recognition and cleavage sequence for signal peptidase I of B. subtilis is Ala-Ala-Ala Ala. The results also indicate that the secretion of the α-amylase in B. subtilis is in accordance with the post-translational transportation mode.     

Kinetic studies of methanol synthesis from CO_2+H_2 over copper based catalysts

The catalytic activity for the synthesis of methanol from carbon dioxide and hydrogen wasmeasured on various binary and ternary catalysts containing copper oxide under a pressure of 10 atm.Among these samples the catalysts, CuO/ZnO/γ-Al_2O_3, demonstrated the highest activity andselectivity to methanol; MnO, as third component, had no promotional effect on the activity of meth-anol formation. Based on a simple power rate law the apparent activation energy estimation and par-tial pressure dependence measurement were accomplished over eight catalysts. The activation energiesvaried from 40 to 120 kJ / mol depending on the composition of catalysts. The rates of methanol for-mation to be 0.3 -- 0.9 order in H_2 and about 0.1 -- 0.2 order in CO_2 were reported.     

Synthesis of porous hematite nanorods loaded with CuO nanocrystals as catalysts for CO oxidation

Porous hematite (α-Fe2O3) nanorods with the diameter of 20-40 nm and the length of 80-300 nm were synthesized by a simple surfactant-assisted method in the presence of cetyltrimethylammonium bromide (CTAB).The α-Fe2O3 nanorods possess a mesostructure with a pore size distribution in the range of 5-12 nm and high surface area,exhibiting high catalytic activity for CO oxidation.CuO nanocrystals were loaded on the surface of porous α-Fe2O3 nanorods by a deposition-precipitation method,and the catalysts exhibited superior activity for catalytic oxidation of CO,as compared with commercial α-Fe2O3 powders supported CuO catalyst.The enhanced catalytic activity was attributed to the strong interaction between the CuO nanocrystals and the support of porous α-Fe2O3 nanorods.     

α-FACTOR-DIRECTED SYNTHESIS AND SECRETION OF β-SUBUNIT OF HUMAN CHORIOGONADOTROPIN (β-hCG)IN YEAST S. cerevisiae

A couple of sbuttle vectors (pYNA and pYNB), containing a sequence coding for the leader regionof the α-factor precursor (α-factor leader sequence) and the origin of 2μm circle, were constructed. ThecDNA for the β-hCG (145aa) was fused in phase to the 3'-end of the α-factor leader sequence andtransformed into the yeast Saccharomyces cercvisiae. The yeast cells carrying this hybrid gene synthesizedand secreted a protein with the immunoactivity of β-hCG in an amount of 45μg per liter of culture. Thepreliminary characterization of this protein is also reported in this paper.     

Utilization of α-olefins obtained by pyrolysis of waste high density polyethylene to synthesize α-olefin-succinic-anhydride based cold flow improvers

A new route of utilization of α-olefin rich hydrocarbon fractions obtained by waste polymer pyrolysis was investigated. α-olefin-succinic-anhydride intermediate-based pour point depressant additives for diesel fuel were synthesized, in which reactions needed α-olefins were obtained by pyrolysis of waste high-density polyethylene (HDPE). Fraction of α-olefins was produced by the de-polymerization of plastic waste in a tube reactor at 500℃ in the absence of catalysts and air. C17～22 range of mixtures of olefins and paraffins were separated for synthesis and then, these hydrocarbons were reacted with maleic-anhydride (MA) for formation of α-olefin-succinic-anhydride intermediates. The olefin-rich hydrocarbon fraction contained approximately 60% of olefins, including 90%～95% α-olefins. Other intermediates were produced in the same way by using commercial C20 α-olefin instead of C17～22 olefin mixture. The two different experimental intermediates with number average molecular weights of 1850g/mol and 1760g/mol were reacted with different alcohols: 1-butanol, 1-hexanol, 1-octanol, i-butanol, and c-hexanol to produce their ester derivatives. The synthesized ten experimental pour point depressants were added in different concentrations to conventional diesel fuel, which had no other additive content before. The structure and efficiency of experimental additives were followed by different standardized and non-standardized methods. Results showed that the experimental additives on the basis of the product of waste pyrolysis were able to decrease not only the pour but also the cloud point and cold filter plugging point (CFPP) of diesel fuel, whose effects could be observed even if the concentration of additives was low. Furthermore, all additives had anti-wear and anti-friction effects in diesel fuel.     

The Concentration of Hydrogen Peroxide Generated during Aggregation of α-Synuclein in vitro Is Lower than 5 nmol／L

Using a fluorometric method with a detection limit of 5 nmol/L, here it is reported that albeit positive results were got from bovine serum albumin (BSA) and chicken ovalbumin (OVA) as published in literature, no detectable amount of hydrogen peroxide (H2O2) was generated during α-synuclein (α-Syn) aggregation in vitro even in the presence of transition metal ions Cu(Ⅱ) or Fe(Ⅲ). The results suggest that the concentration of H2O2 generated during aggregation of α-Syn in vitro be lower than 5 nmol/L beyond the detection limit of the adopted method and it is far too poor to be responsible for the cytotoxicity of α-Syn aggregates, thus allowing people to extensively elucidate the mechanism underlying neurotoxicifies of the aggregates formed by some amyloidogenic proteins.     

CLONING AND ANALYSIS OF HISTONE 3 GENE AND RUBISCO SMALL SUBUNIT GENE OF RICE

We have isolated and determined the DNA sequence of several genes from the nucleus of rice (Oryza sativa, IR26). We screened a genomic library of rice IR26 and isolated a 14.8 kb segment containing an H3 gene and an H3-like pseudogene. Sequence analysis showed that the coding sequence of the rice H3 gene is 405 bp in length, and the 5' and 3' noncoding regions contain several regulatory sequences common to eukaryotic or histone genes. The codon usage of the rice H3 gene is highly unusual in that the third codon position is 98% G and C. Southern blotting analysis suggested that the copy number of the H3 gene is around 50 per diploid rice genuine. From the same rice geaomic library, we have identified rbcS, which codes for the small subunit of ribulose-1, 5-bisphosphate carboxylase-oxygenase (Rubisco). The rbcS sequence is interrupted by an intron at the same location in both rice and wheat. The first 18 amino acids of the transit peptide in rice and wheat rbcS are identical     

In vivo investigation of the role of SfmO2 in saframycin A biosynthesis by structural characterization of the analogue saframycin O

Saframycin A(SFM-A),a tetrahydroisoquinoline antibiotic isolated from Streptomyces lavendulae,shows potent anti-proliferation activities against a variety of tumor cell lines,and shares the core structure with ecteinascidin 743(ET-743),the anticancer drug for soft-tissue sarcoma.Characterization of the SFM-A biosynthetic gene cluster revealed three nonribosomal peptide synthetase genes and a series of genes encoding oxygenases.To investigate the function of sfmO2 gene,encoding a FAD-dependent monooxygenase/hydroxylase,we constructed the gene replacement mutant(△sfmO2) strain S.lavendulae TL2007 and the corresponding gene complementation mutant strain S.lavendulae TL2008.A novel compound,SFM-O,was isolated from the △sfmO2 replacement mutant strain and its structure was characterized by comparison to the HRMS and NMR spectra of SFM-A.These findings indicated that SfmO2 is responsible for the oxidation of ring A in the biosynthetic pathway of SFM-A,and the new compound SFM-O could be considered as an advanced intermediate in the semisynthesis of ET-743.     

KINETICS OF THE ZIEGLER-NATTA POLYMERIZATION OF 1-OCTENE(I)----THE KINETICS OF THE STATIONARY PERIOD

The kinetics of the Ziegler-Natta polymerization of C6~C20 α-defines has been seldom studied,which showed no apparent differences from the kinetics of the polymerization of ethylene and propylene.1-Octene polymerizes in alkanes in a solution state,differing from the slurry polymerization of ethylene.We have found and studied the inordinary kinetics of the polymerization of 1-octene on α-TiCl3-AlEt3[or-Al(i-Bu)3,-AlEt2Cl] catalysts in n-heptane,and have given a primary explanation to this phenomenon.     

Influence of PEG 6000 on gallium oxide（Ga2O3） polymorphs and photocatalytic properties

Three different nanorod-like gallium oxides with mono/poly-crystalline nature(α, β, and α/β-Ga2O3) were prepared by regulating the amount of polyethylene glycol(PEG) 6000 in the range of 0.2–0.8 g proportionally via a hydrothermal method combined with further calcination. The bandgap of the products, given by UV-Vis diffuse reflectance spectra(UV-Vis DRS), was in the order of α-Ga2O3 > α/β-Ga2O3 > β-Ga2O3. To further investigate the photocatalysis performance of the catalysts, the decomposition of rhodamine B(Rh B) by Ga2O3 under UV light illumination(λ < 387 nm) was presented and complete degradation could be achieved within 30 min, a result that showed the highest efficiency. The photocatalytic oxidation mechanism is further discussed and prominently related to the active species: hydroxyl radical(·OH) and superoxide radical(O·-2), which were confirmed by electron paramagnetic resonance(EPR).     

Stress-induced Solid-Solid Crystal Transition in Trans-1,4-polyisoprene

The polymorphic transition of trans-1,4-polyisoprene(TPI) during stretching was investigated by in situ wide-angle X-ray diffraction and Fourier transform infrared spectroscopy. The influences of the initial structure, stretching temperature, and strain rate on the contents of different crystal modifications(α, β) were explored. The results confirm that the α-β transition occurs during stretching of TPI that only contains αcrystal(α-TPI). When the stress is relaxed, the β crystal formed during stretching remains, which indicates that the transition is irreversible. On the other hand, stretching of TPI that only contains β crystal(β-TPI) results in orientated β crystal. No β-α transition occurs during stretching. The different structures of stretched α-TPI and β-TPI exclude the previously proposed "melting-recrystallization mechanism". The α-β transition depends significantly on temperature and strain rate, indicating the transition is governed both by thermodynamics and kinetics. Our results support a solid-solid transition mechanism rather than a melting-recrystallization mechanism. The irreversible nature of the transition is attributed to the metastability of the β phase in the unstretched state. Different from the "β phases" that appear in polymers with stress-induced reversible transitions, e.g. poly(butylene terephthalate) and poly(butylene succinate), the stability of β phase in TPI is high that can be long-lived.The strain rate dependence of α-β transition hinders the determination of critical stress for the transition. It further indicates that the local stress within the sample is more heterogeneous at higher strain rates.     

Isolation and Characterization of Two Novel Gene Encoded Alkaline Esterases from an Alkaline Soil Metagenomic Library

<正>The development of industrial biotechnology has created an increasing demand for alkaline lipolytic enzymes with functional diversity.In this study,an alkaline soil metagenomic library was constructed to search for new lipolytic enzymes.Two novel gene encoded alkaline esterases(designated as estA and estB) were isolated by functional screening from the library.The estA gene consisted of 834 bp and coded for 277 amino acids with a molecular mass of 29998.Amino acid sequence homology analysis indicates that EstA belongs toα/βhydrolase fold family 4.4(abH4.4),with EstA being the smallest member of that family yet reported.The estB gene consisted of 1185 bp and encoded 394 amino acids with a theoretical molecular mass of 40090.Its conserved domain analysis shows that EstB belongs to the GDSL hydrolase superfamily.Both EstA and EstB exhibit only moderate identity(38%) in amino acid sequence to the known lipolytic enzyme genes in the database.The two genes were respectively expressed in Escherichia coli and the protein products were purified for functional characterization.While the expressed EstA did not exhibit the functional properties that were superior to those of other esterases previously reported,the EstB was stable at temperature up to 45℃and its maximum activity was measured to be 53.6 U/mg at pH=10.Both the enzymes have further enriched the diversity of the lipolytic enzymes database and also appear to be promising biocatalysts for potential biotechnological application.