Discovery and characterization of a novel poly-mannuronate preferred alginate lyase: The first member of a new polysaccharide lyase family.

Alginate is one of the most important marine colloidal polysaccharides, and its oligosaccharides have been proven to possess diverse biological functions. Alginate lyases could specifically degrade alginate and therefore serve as desirable tools for the research and development of alginate. In this report, a novel catalytic domain, which demonstrated no significant sequence similarity with all previously defined functional domains, was verified to exhibit a random endo-acting lyase activity to alginate. The action pattern analysis revealed that the heterologously expressed protein, named Aly44A, preferred to degrade polyM. Its minimum substrates and the minimum products were identified as unsaturated alginate trisaccharides and disaccharides, respectively. Based on the sequence novelty of Aly44A and its homologs, a new polysaccharide lyase family (PL44) was proposed. The discovery of the novel enzyme and polysaccharide lyase family provided a new entrance for the gene-mining and acquiring of alginate lyases, and would facilitate to the utilization of alginate and its oligosaccharides.

Effect of Au Plasmonic Material on Poly M-Toluidine for Photoelectrochemical Hydrogen Generation from Sewage Water.

This study provides H2 gas as a renewable energy source from sewage water splitting reaction using a PMT/Au photocathode. So, this study has a dual benefit for hydrogen generation; at the same time, it removes the contaminations of sewage water. The preparation of the PMT is carried out through the polymerization process from an acid medium. Then, the Au sputter was carried out using the sputter device under different times (1 and 2 min) for PMT/Au-1 min and PMT/Au-2min, respectively. The complete analyses confirm the chemical structure, such as XRD, FTIR, HNMR, SEM, and Vis-UV optical analyses. The prepared electrode PMT/Au is used for the hydrogen generation reaction using Na2S2O3 or sewage water as an electrolyte. The PMT crystalline size is 15 nm. The incident photon to current efficiency (IPCE) efficiency increases from 2.3 to 3.6% (at 390 nm), and the number of H2 moles increases from 8.4 to 33.1 mmol h-1 cm-2 for using Na2S2O3 and sewage water as electrolyte, respectively. Moreover, all the thermodynamic parameters, such as activation energy (Ea), enthalpy (ΔH*), and entropy (ΔS*), were calculated; additionally, a simple mechanism is mentioned for the water-splitting reaction.

Identification and Characterization of a New Cold-Adapted and Alkaline Alginate Lyase TsAly7A from Thalassomonas sp. LD5 Produces Alginate Oligosaccharides with High Degree of Polymerization.

Alginate oligosaccharides (AOS) and their derivatives become popular due to their favorable biological activity, and the key to producing functional AOS is to find efficient alginate lyases. This study showed one alginate lyase TsAly7A found in Thalassomonas sp. LD5, which was predicted to have excellent industrial properties. Bioinformatics analysis and enzymatic properties of recombinant TsAly7A (rTsAly7A) were investigated. TsAly7A belonged to the fifth subfamily of polysaccharide lyase family 7 (PL7). The optimal temperature and pH of rTsAly7A was 30 °C and 9.1 in Glycine-NaOH buffer, respectively. The pH stability of rTsAly7A under alkaline conditions was pretty good and it can remain at above 90% of the initial activity at pH 8.9 in Glycine-NaOH buffer for 12 h. In the presence of 100 mM NaCl, rTsAly7A showed the highest activity, while in the absence of NaCl, 50% of the highest activity was observed. The rTsAly7A was an endo-type alginate lyase, and its end-products of alginate degradation were unsaturated oligosaccharides (degree of polymerization 2-6). Collectively, the rTsAly7A may be a good industrial production tool for producing AOS with high degree of polymerization.

Production, Characterization, and Application of an Alginate Lyase, AMOR_PL7A, from Hot Vents in the Arctic Mid-Ocean Ridge.

Enzymatic depolymerization of seaweed polysaccharides is gaining interest for the production of functional oligosaccharides and fermentable sugars. We describe a thermostable alginate lyase belonging to Polysaccharide Lyase family 7 (PL7), which can be used to degrade brown seaweed, Saccharina latissima, at conditions also suitable for a commercial cellulase cocktail (Cellic CTec2). This enzyme, AMOR_PL7A, is a ß-d-mannuronate specific (EC 4.2.2.3) endoacting alginate lyase, which degrades alginate and poly mannuronate within a broad range of pH, temperature and salinity. At 65 °C and pH 6.0, its Km and kcat values for sodium alginate are 0.51 ± 0.09 mg/mL and 7.8 ± 0.3 s-1 respectively. Degradation of seaweed with blends of Cellic CTec2 and AMOR_PL7A at 55 °C in seawater showed that the lyase efficiently reduces viscosity and increases glucose solublization. Thus, AMOR_PL7A may be useful in development of efficient protocols for enzymatic seaweed processing.

Development of effective nano-biosorbent based on poly m-phenylenediamine grafted dextrin for removal of Pb (II) and methylene blue from water.

Novel nano-biosorbent was successfully prepared based on dextrin-g-poly m-phenylenediamine (DgPmPDA) and graphene oxide (GO) by using the solution blending technique and was found to be an effective adsorbent for Pb(II) and methylene blue (MB) dye. The nano-biosorbent was characterized by FTIR, CHN, XRD, SEM, AFM and TGA. The adsorption process depended on the pH of solution, adsorbent dosage, contact time, initial concentration of Pb(II) and MB, and temperature. Experimental results were in good agreement with Langmuir isotherm model for both Pb(II) and MB adsorption. The maximum adsorption capacity (Qmax) of Pb(II) and MB was found to be 80 mg/g and 76.33 mg/g, respectively, at optimum conditions. Results showed that the kinetics of Pb(II) and MB onto the DgPmPDA@GO nano-biosorbent followed the pseudo-first-order and pseudo-second-order models, respectively. Thermodynamic parameters exhibited the endothermic and spontaneous nature of the sorption process. A possible mechanism of adsorption was suggested where π-π stacking interactions, H-bonding interaction and electrostatic attraction controlled the MB adsorption and chelation and electrostatic attraction controlled the Pb(II) adsorption.

Characterization of a Novel PolyM-Preferred Alginate Lyase from Marine Vibrio splendidus OU02.

Alginate lyases are enzymes that degrade alginate into oligosaccharides which possess a variety of biological activities. Discovering and characterizing novel alginate lyases has great significance for industrial and medical applications. In this study, we reported a novel alginate lyase, AlyA-OU02, derived from the marine Vibrio splendidus OU02. The BLASTP searches showed that AlyA-OU02 belonged to polysaccharide lyase family 7 (PL7) and contained two consecutive PL7 domains, which was rare among the alginate lyases in PL7 family. Both the two domains, AlyAa and AlyAb, had lyase activities, while AlyAa exhibited polyM preference, and AlyAb was polyG-preferred. In addition, the enzyme activity of AlyAa was much higher than AlyAb at 25 °C. The full-length enzyme of AlyA-OU02 showed polyM preference, which was the same as AlyAa. AlyAa degraded alginate into di-, tri-, and tetra-alginate oligosaccharides, while AlyAb degraded alginate into tri-, tetra-, and penta-alginate oligosaccharides. The degraded products of AlyA-OU02 were similar to AlyAa. Our work provided a potential candidate in the application of alginate oligosaccharide production and the characterization of the two domains might provide insights into the use of alginate of this organism.

In Silico Tracking of Individual Species Accelerating Progress in Macromolecular Engineering and Design.

The beneficial use of computer simulations to track the microstructural evolution of individual species is highlighted in view of macromolecular engineering and design, considering two case studies on catalytic polymerization, and both "low" (<100) and "high" (>100) chain lengths, that is, i) atom transfer radical copolymerization of n-butyl acrylate and styrene aiming at the synthesis of functional macrospecies of "identical' chain length; and ii) chain shuttling polymerization of ethylene and 1-octene toward the production of segmented block copolymers with "soft" and "hard" segments. Model parameters are validated and/or tuned based on literature data. The modeling strategy supports the future identification of chemical structure-polymer property relationships and is based on the combination of principles from polymer reaction engineering, chemistry, and physics.

Biochemical Characterization and Degradation Pattern of a Unique pH-Stable PolyM-Specific Alginate Lyase from Newly Isolated Serratia marcescens NJ-07.

Enzymatic preparation of alginate oligosaccharides with versatile bioactivities by alginate lyases has attracted increasing attention due to its featured characteristics, such as wild condition and specific products. In this study, AlgNJ-07, a novel polyM-specific alginate lyase with high specific activity and pH stability, has been purified from the newly isolated marine bacterium Serratia marcescens NJ-07. It has a molecular weight of approximately 25 kDa and exhibits the maximal activity of 2742.5 U/mg towards sodium alginate under 40 °C at pH 9.0. Additionally, AlgNJ-07 could retain more than 95% of its activity at pH range of 8.0-10.0, indicating it possesses excellent pH-stability. Moreover, it shows high activity and affinity towards polyM block and no activity to polyG block, which suggests that it is a strict polyM-specific alginate lyase. The degradation pattern of AlgNJ-07 has also been explored. The activity of AlgNJ-07 could be activated by NaCl with a low concentration (100-300 mM). It can be observed that AlgNJ-07 can recognize the trisaccharide as the minimal substrate and hydrolyze the trisaccharide into monosaccharide and disaccharide. The TLC and ESI-MS analysis indicate that it can hydrolyze substrates in a unique endolytic manner, producing not only oligosaccharides with Dp of 2-5 but also a large fraction of monosaccharide. Therefore, it may be a potent tool to produce alginate oligosaccharides with lower Dps (degree of polymerization).

Correlation between Genetic Variants and Polymorphism of Caveolin and Sudden Unexplained Death / 法医学杂志

Objective T o explore the genetic variation sites of caveolin (C A V ) and their correlation w ith sudden unexplained death (SU D ).Methods The blood sam ples w ere collected from SU D group (71 cases), coronary artery disease (C A D ) group (62 cases) and control group (60 cases), respectively. T he genom e D N A w ere extracted and sequencing w as perform ed directly by am plifying gene coding region and exon-intron splicing region of CAV1 and CAV3 using PC R . T he type of heritable variation of CVA w as con-firm ed and statistical analysis w as perform ed. Results A total of 4 variation sites that m aybe significa-tive w ere identified in SU D group, and tw o w ere new found w hich w ere CAV1: c.45C>T (T 15T ) and CAV1:c.512G>A (R 171H ), and tw o w ere SN P loci w hich w ere CAV1:c.246C>T (rs35242077) and CAV3:c.99C>T (rs1008642) and had significant difference (P<0.05) in allele and genotype frequencies betw een SU D and control groups. Forem entioned variation sites w ere not found in C A D group. Conclu-sion T he variants of CAV1 and CAV3 m ay be correlated w ith a part of SU D group.

Sixty-six Biallelic Genetic Markers on Y chromosome by MALDI-TOF-MS / 法医学杂志

Objective T o analyse the genetic polym orphism s of 66 biallelic genetic m arkers on Y chro-m osom e in E astern C hinese H an population, and evaluate their values in forensic application. Methods G enotyping of 66 biallelic genetic m arkers on Y chrom osom e w as studied in 205 unrelated m ales of E astern C hinese H an population by m ultiplex PC R com bined m atrix-assisted laser desorption/ionization tim e-of-flight m ass spectrom etry (M A L D I-T O F-M S ). T he allele frequencies on the loci to be tested w ere calculated by direct counting m ethod, and the gene diversity (G D ) and haplotype diversity (H D ) w ere calculated by corresponding form ulas. T he haplotypes of this system w ere tested by softw are A rlequin v3.5.2.2 and the com parison of population genetics w ere analyzed. Results A total of 60 biallelic genetic m arkers on Y chrom osom e w ere polym orphic in m ales of E astern C hinese H an population, and the ranges of G D w ere from 0.0385 to 0.5019. E ighty-five different haplotypes w ere observed and the H D w as 0.9703. T he differences of partial SN P loci betw een the H an population of E astern C hina and that of X injiang and G uangdong w ere statistically significance. Conclusion Sixty biallelic genetic m arkers and the detection system can com plem entally provide genetic inform ation in kinship testing and individual identification. T he M A L D I-T O F-M S technology is able to type biallelic genetic m arkers.

One-pot inimer promoted ROCP synthesis of branched copolyesters using α-hydroxy-γ-butyrolactone as the branching reagent.

An array of branched poly(ɛ-caprolactone)s was successfully synthesized using an one-pot inimer promoted ring-opening multibranching copolymerization (ROCP) reaction. The biorenewable, commercially available yet unexploited comonomer and initiator 2-hydroxy-γ-butyrolactone was chosen as the inimer to extend the use of 5-membered lactones to branched structures and simultaneously avoiding the typical tedious work involved in the inimer preparation. Reactions were carried out both in bulk and in solution using stannous octoate (Sn(Oct)2) as the catalyst. Polymerizations with inimer equivalents varying from 0.01 to 0.2 were conducted which resulted in polymers with a degree of branching ranging from 0.049 to 0.124. Detailed ROCP kinetics of different inimer systems were compared to illustrate the branch formation mechanism. The resulting polymer structures were confirmed by 1H, 13C, and 1H-13C HSQC NMR and SEC (RI detector and triple detectors). The thermal properties of polymers with different degree of branching were investigated by DSC, confirming the branch formation. Through this work, we have extended the current use of the non-homopolymerizable γ-butyrolactone to the branched polymers and thoroughly examined its behaviors in ROCP. © 2016 The Authors. Journal of Polymer Science Part A: Polymer Chemistry Published by Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2016, 54, 1908-1918.

Well-controlled ATRP of 2-(2-(2-Azidoethyoxy)ethoxy)ethyl Methacrylate for High-density Click Functionalization of Polymers and Metallic Substrates.

The combination of atom transfer radical polymerization (ATRP) and click chemistry has created unprecedented opportunities for controlled syntheses of functional polymers. ATRP of azido-bearing methacrylate monomers (e.g. 2-(2-(2-azidoethyoxy)ethoxy)ethyl methacrylate, AzTEGMA), however, proceeded with poor control at commonly adopted temperature of 50 °C, resulting in significant side reactions. By lowering reaction temperature and monomer concentrations, well-defined pAzTEGMA with significantly reduced polydispersity were prepared within a reasonable timeframe. Upon subsequent functionalization of the side chains of pAzTEGMA via Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC) click chemistry, functional polymers with number-average molecular weights (Mn) up to 22 kDa with narrow polydispersity (PDI < 1.30) were obtained. Applying the optimized polymerization condition, we also grafted pAzTEGMA brushes from Ti6Al4 substrates by surface-initiated ATRP (SI-ATRP), and effectively functionalized the azide-terminated side chains with hydrophobic and hydrophilic alkynes by CuAAC. The well-controlled ATRP of azido-bearing methacrylates and subsequent facile high-density functionalization of the side chains of the polymethacrylates via CuAAC offers a useful tool for engineering functional polymers or surfaces for diverse applications.

Photoinduced Acrylate Polymerization: Unexpected Reduction in Chain Branching.

The branching stemming from midchain radical formation in n-butyl acrylate polymerization is investigated via melt-state (13) C NMR measurements. The dependence of the degree of branching (DB) on the monomer conversion of the system is examined for photoinduced polymerizations, revealing a steady increase in branching with conversion. For polymerization at moderate light intensities, an increase in branching from 0.03% to 0.37% is observed for polymerizations at 60 °C, which is fivefold below the level of branching observed in thermally initiated polymerizations under otherwise identical reaction conditions. The reason for this overall reduction in branching remains momentarily unclear; yet, a strong dependence of branching on light intensity is observed. While polymerization under a 1 W LED lamp results at almost full monomer conversion in branching degrees of 0.22%, polymerization under a 400 W lamp yields 1.81% of chain branches.

Forensic Investigation of GoldeneyeTM DNA ID 22NC Kit / 法医学杂志

Objective To investigate the genetic data of 21 autosom al STR included in G oldeneyeTM DNA ID 22N CK it in Chinese H an nationality and to evaluate the forensic application. Methods B y detected 500 unrelated healthy individuals in Chinese H an nationality of East China w ith G oldeneyeTM DNA ID 22N CK it, allele frequencies, population genetics param eters and linkage disequilibrium inform ation of the 21 autosom al STR w ere statistically analyzed. Results In the 21 autosom al STR , no deviations from H ardy-W einberg equilibrium w ere detected and all loci w ere independent form each other. D P values of 21 au-tosom al STR w ere all above 0.85, and the com bined discrim ination pow er w as 1-3.616 5×10-26. Com bined m ean exclusion chance of this system in duo cases w as 1-2.786 81×10-6, in trio cases w as 1-8.545 82× 10-10. Conclusion Tw enty-one autosom al STR included in G oldeneyeTM DNA ID 22N CK it are highly polym orphic in the H an nationality. Com bined w ith G oldeneyeTM DNA ID 20A K it, the kit can satisfy the needs for full-sibling testing and facilitate the solution of this kind of case tools.

Validation of GlobalFiler?PCR Amplification Kit and the STR Polymorphism / 法医学杂志

Objective To test the technical param eters of GlobalFiler?PC R A m plification K it for its ap-plication to forensic application value and to investigate the genetic polym orphism s. Methods The valida-tion w as conducted in sensitivity, m ixed sam ples, species specificity, adaptability, survivability, consistency, peak height balance and stability. The am plification and detection of the genom ic D N A from 373 unre-lated individuals from B eijing H an nationality w ere extracted by autom ation w orkstation. Results Global-Filer?PC R A m plification K it w as adaptive to som e m ixed, degraded and inhibited sam ples. The pow er of sensitivity and adaptability and peak height balance show ed w ell. The distributions of genotype fre-quencies for 21 STR loci in the population w ere all in accordance w ith H ardy-W einberg equilibrium (P>0.05). The PIC value of the 21 STR loci w as am ong 0.536 to 0.940; the H value w as am ong 0.558 to 0.933; the D P value w as am ong 0.783 to 0.992; the PE value w as am ong 0.243 to 0.874. Conclusion GlobalFiler?PC R A m plification K it is suitable for crim inal cases and D N A database in forensic practice. A nd 21 STR loci in B eijing H an nationality have high polym orphism , w hich have ap-plication value in forensic practice and population genetics.

Genetic Polymorphisms of 26 Y-STR Loci in Fujian She Nationality and Its Forensic Application / 法医学杂志

Objective To study the forensic application of G oldeneyeTM D N A ID 26Y K it in the She na-tionality. Methods Through capillary electrophoresis, the genotype of 26 Y-STR loci were analyzed in 53 unrelated m ale individuals from Fujian She nationality. The population genetics param eters such as allele frequency and haplotype diversity were calculated. The com parisons am ong the She nationality and the other nationalities were analyzed. Results A total of 126 alleles were observed on the 26 Y-STR loci of 53 unrelated m ale individuals. The allele frequencies and G D value ranged from 0.010 1 to 0.886 8 and 0.211 2 to 0.846 2, respectively. The G D value w as greater than 0.5 in the 19 loci. A total of 47 hap-lotypes were observed. B ased on RST, m ultidim ensional scaling plot indicated that the genetic relationship am ong Fujian She nationality and M innan H an nationality w as closest, follow ed by Southern C hina H an nationality and N orthern C hina nationality. Conclusion G oldeneyeTM D N A ID 26Y K it including 26 Y-STR loci has good polym orphism in the She nationality. A s an additional system , it has forensic appli-cation value in som e special cases.

Genetic Polymorphisms of 21 Autosomal STR Loci of Fujian Han Population / 法医学杂志

Objective To investigate the genetic polym orphism s of 21 autosom al STR loci of Fujian H an population and evaluate the forensic application value of GlobalFilerTM E xpress kit. Methods A m plified w ith GlobalFilerTM E xpress kit, DNA sam ples were obtained from 741 unrelated individuals of Fujian H an population. The population genetics param eters of 21 autosom al STR loci were calculated. Results The 21 autosom al STR loci were found to be no deviation from Hardy-Weinberg equilibration (P>0.05) and relatively abundant in high polym orphism . H eterozygosity ranged from 0.589 to 0.914, pow er of dis-crim ination ranged from 0.754 to 0.992, polym orphic inform ation content ranged from 0.520 to 0.940, and pow er of exclusion ranged from 0.278 to 0.825. The SE33 locus was the highest degree in poly-m orphism . Conclusion The 21 STR loci of GlobalFilerTM E xpress kit have high value in discrim ination pow er and can be useful in personal identification and paternity test in Fujian H an population.

Progress in Association between Genetic Correlation and Human Violent Behavior / 法医学杂志

H um an violent behavior is a com plex behavior w hich is influenced by genetic and environ-m ental factors. T here is a trend in investigating the m echanism of violent behavior by using the genetic m ethods. T his article review s several candidate genes and advances in epigenetics w hich are associated w ith violent behavior. T he prospects and significance of violent behavior research from the view of gene polym orphism and epigenetics are also discussed.

Expression of SFRP5 mRNA in Rat Skeletal Muscle after Contusion / 法医学杂志

Objective T o investigate the relationship betw een the expression of secreted frizzled-related protein 5 (SFR P5) m RNA and the tim e interval after skeletal m uscle injury in rats by real-tim e PC R . Methods A total of ninety SD rats w ere random ly divided into the contusion groups at different tim es including 4 h, 8 h, 12 h, 16 h, 20 h, 24 h, 28 h, 32 h, 36 h, 40 h, 44 h, 48 h after contusion, incision groups at different tim es including 4h and 8h after incision and the control group. T he sam ples w ere taken from the contused zone at different tim e points. T he total RNA w as isolated from the sam ples and re-versely transcribed to analyze the expression levels of SFRP5 m RNA . Results C om pared to the control group, the expression of SFRP5 m RNA in contusion groups w ere dow n-regulated w ithin 48 h after con-tusion and reached the low est level at 20 h, and the expression of SFRP5 m RNA gradually increased from 20 h to 48 h after contusion. T he expression of SFRP5 m RNA in the incised groups w ere signifi-cantly low er than that of the contusion groups at 4 h after injury. A t the tim e of 8 h, the expression levels betw een the contusion and incision groups show ed no statistically significant difference. Conclusion It is suggested that SFRP5 m RNA analysis m ay show regular expression and can be a m arker for estim ation of skeletal m uscle injury age.

Establishing a 29 Y-STR Loci Multiplex PCR System / 法医学杂志

Objective To establish a 29 Y-STRloci multiplex PC Rsystemfor investigating the genetic polymorphisms and to assess its application value in forensic science. Methods Amultiplex PC Rsystemw as established using a five color fluorescence labeling 29 Y-STRloci (DYS456, DYS389Ⅰ, DYS437, DYS447, DYS389Ⅱ, DYS438, DYS522, DYS460, DYS458, DYS622, DYS390, DYS392, DYS448, DYS449, DYS391, Y-GATA-H4, DYS388, DYS19, DYS385a/b, DYS527a/b, DYS393, DYS459a/b, DYS635, DYS439, DYS570 and DYS627) for multiple amplification and capillary electrophoresis. And its applicability w as validated w ith genetic polymorphismdata of 29 Y-STRof unrelated 2 000 male samples in Shandong Han population. Results Atotal of 1981different haplotypes of 2 000 individuals show ed genotype diver-sity betw een 0.370 0 and 0.965 4. The systemprovided stable and accurate typing w ith high sensitivity of 0.05 ng. It satisfied the needs of variety of routine biological samples. Conclusion The 29 Y-STRloci multiplex PC Rsystemcould be applied for actual cases and establishment of Y-STRdatabase. In addi-tion, it has great significance in forensic science practices and related research.