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The influences of biologically synthesized gold nanoparticles (bio-GNPs) and ultra thin silver layer (UTSL) on the polymer light emitting diodes (PLEDs) are investigated for the first time. The performance of the fabricated PLEDs is enhanced by embedding of bio-GNPs into the hole transport layer (HTL). Furthermore, the tailored device architecture containing UTSL increases the electron transport through lightning rod effect. Bio-GNPs are successfully produced as spherical shape with size of circa 10.4 nm. Poly[2-methoxy-5-(2-ethylhexyloxy)-1,4-phenylene vinylene (MEH-PPV) and [poly(3,4-ethylenedioxythiophene):poly (styrenesulfonate); (PEDOT:PSS)/bio-GNPs blends are used as an active layer and HTL, respectively. Novel PLEDs fabricated with 0.125 wt% bio-GNPs/PEDOT:PSS and 0.5 nm UTSL exhibit nearly 2.5-fold enhancement in the device efficiency.

Çakır Hatır, Pınar (Arel Author), In this study, poly(3-hydroxybutyrate) (PHB) production from a newly isolated obligate alkaliphilic Bacillus marmarensis DSM 21297 was investigated to evaluate the ability of obligate alkaliphilic strain to produce a biopolymer. Additionally, electrospun nanofibers from B. marmarensis PHB (Bm-PHB) were generated using Bm-PHB/polycaprolactone (PCL) blend to evaluate the applicability of Bm-PHB. According to the experimental results, the metabolic activity of B. marmarensis decreased the pH of the medium by generating H+ ions to initiate Bm-PHB production, which was achieved at pH below 9.0. Regarding medium components, the addition of MgSO4 center dot 7H(2)O and KH2PO4 to the medium containing 1% glucose enhanced the amount of Bm-PHB synthesis, and an approximately 60% increase in PHB concentration was obtained in the presence of mineral salts.Based on FTIR analysis, the chemical structures of Bm-PHB and commercial PHB were found to be highly similar. Additionally, the T-g and T-m values of Bm-PHB were determined to be 17.77 degrees C and 165.17 degrees C, respectively. Moreover, Bm-PHB/PCL composite scaffold was generated by electrospinning method that produced nanofibers between 150 and 400 nm in diameter, with an average of 250 nm. To our knowledge, this is the first report to produce PHB from an obligate alkaliphilic Bacillus strain and PHB scaffold.

Maltooligosaccharides producing amylases are required in the food industry, especially in breadmaking. The Bacillus subtilis strain SDP1 amylase hydrolyses starch to produce maltotriose and maltotetraose along with maltose after prolonged reactions of 5 h. Bacillus subtilis strain SDP1 was isolated from the rhizosphere of Acacia cyanophylla Lindley from the Cukurova region of Turkey. The highest enzyme production was achieved with soluble starch as the carbon and yeast extract as the nitrogen source and at pH 7.0 and 37°C. Under optimized culture conditions, 68.49 U/mL activity was obtained. SDP1 α-amylase had molecular weight of 61 kD. The optimum pH of the enzyme was 7.0 and was highly active at pH ranging from 5.0 to 9.0. The optimum temperature of the crude enzyme was 60°C, and it retained 83% and 74% of its initial activity after 1 h and 2 h incubation periods, respectively, at 50°C. While, Mn+2 has a stimulatory effect on the activity, Ca+2, Mg+2, Na+ did not effect the enzyme activity. Fe+3, Ni+2, ...

In this work, the protein pattern of novel Halomonas smyrnensis AAD6T was compared to that of Halomonas salina DSMZ5928T, which is the closest species on the basis of 16S rRNA sequence, to understand how AAD6T differs from type strains. Using high resolution NEPHEGE technique, the whole cell protein composition patterns of both Halomonas salina DSMZ5928T and H. smyrnensis AAD6T were mapped. The expressed proteins of the two microorganisms were mostly located at the acidic side of the gels, at molecular weight values of 60 to 17 kDa, and at isoelectric points 3.8 to 6.0, where they share a significant number of common protein spots. Identification and characterization of protein spots via whole genome sequencing data indicated that these two microorganisms used similar pathways, especially TCA cycle, for their survival; in other words, for their energy requirements. On the other hand, the protein expression differences in AAD6T and H. salina DSMZ 5928T showed that they prefer different metabolic pathways for lipid biosynthesis and in adaptation to extreme environments. Thus, we suggested that phylogenetic dissimilarities between these microorganisms could be related to the protein expression differences; in other words, metabolic flux differences in AAD6T and H. salina DSMZ 5928T. This is the first study to explain the dissimilarities of phenotypic characters and DNA-DNA hybridization between type strain and novel strain AAD6T by using protein expression differences.

This paper reports the isolation and identification of chitinase-producing Bacillus from chitin-containing wastes, production of a thermostable and alkaline chitinasese, and enzyme characterization. Bacillus thuringiensis subsp. kurstaki HBK-51 was isolated from soil and was identified. Chitinase was obtained from supernatant of B. thuringiensis HBK-51 strain and showed its optimum activity at 110°C and at pH 9.0. Following 3 hours of incubation period, the enzyme showed a high level of activity at 110°C (96% remaining activity) and between pH 9.0 and 12.0 (98% remaining activity). Considering these characteristics, the enzyme was described as hyperthermophile-thermostable and highly alkaline. Two bands of the enzyme weighing 50 and 125 kDa were obtained following 12% SDS-PAGE analyses. Among the metal ions and chemicals used, Ni2+ (32%), K+ (44%), and Cu2+ (56%) increased the enzyme activity while EDTA (7%), SDS (7%), Hg2+ (11%), and ethyl-acetimidate (20%) decreased the activity of the enzyme. Bacillus thuringiensis subsp. kurstaki HBK-51 is an important strain which can be used in several biotechnological applications as a chitinase producer.

Alkaline proteases are one of the most important group of enzymes that are indispensable in a number of different industrial sectors. In this work, the effect of copper ions (Cu2+) was investigated for improving the thermostability and hydrolytic performance of Bacillus clausii GMBE 42 alkaline protease at different temperatures (45–65°C). Maximal residual activity was observed in the presence of 5 mM CuCl2. The enzyme was thermoinactivated according to first-order kinetics. A stabilization effect caused by copper ions was the result of a decrease in both autolysis and thermoinactivation rates. Thermodynamic analysis of the thermoinactivation process showed that Ea,i, ΔGi, and ΔHi values of the enzyme were higher in the presence of copper ions, but there was no measurable change in ΔSi values. These results show the thermostabilizing potential of copper ions on the enzyme. Lower Km values and higher kcat and kcat/Km values were obtained in the presence of copper ions, which is an indication of the nonessential activation of the enzyme by copper ions. Thermodynamic analysis of casein hydrolysis showed that in presence of copper ions Ea, ΔG≠, ΔH≠, , and values of enzyme were lower, but there was no change in ΔS≠ values. This is so far the first study that investigates the effect of cations on the basic catalytic and thermodynamic properties of an alkaline serine protease, which may be used to remove protein wastes from various industries such as food and leather processing.

Proteases are largely employed in biocatalysis. In order to increase the number of their applications it is useful to shed light on the reasons that cause a non-optimal activity of these enzymes when used in inactivating experimental conditions (e.g., in the presence of co-solvent to favor substrate dissolution). To this end the effect of different mono alcohols on the activity and the conformation of alkaline protease from Bacillus clausii GMBAE 42 was investigated. We found that the enzyme in the presence of 20–25% of methanol, ethanol, 1-propanol or 2-propanol halves its activity. At the concentration of 10%, all the alcohols caused a slightly more intense far-UV (CD) circular dichroism signal of the protease at around 208 and 220 nm with respect to the protein in only buffer, which suggests an increase of helicity in the secondary structure of the protein. Monitoring the shift of the fluorescence emission of the protease with respect to that of the standard N -acetyl- l -tryptophan-ethyl ester, we suggest that with all the alcohols tested the decrease of activity might be due to the loss of tertiary structure (even though at a lower extents in methanol and ethanol compared to 1-propanol and 2-propanol).

An alkali tolerant Bacillus strain having extracellular serine alkaline protease activity was newly isolated from compost and identified as Bacillus clausii GMBE 22. An alkaline protease (AP22) was 4.66-fold purified in 51.5% yield from Bacillus clausii GMBE 22 by ethanol precipitation and DEAE-cellulose anion exchange chromatography. The purified enzyme was identified as serine protease by LC-ESI-MS analysis. Its complete inhibition by phenylmethanesulfonylfluoride (PMSF) also justified that it is a serine alkaline protease. The molecular weight of the enzyme is 25.4 kDa. Optimal temperature and pH values are 60 degrees C and 12.0, respectively. The enzyme showed highest specificity to N-Suc-Ala-Ala-Pro-Phe-pNA. The K(m) and k(cat) values for hydrolysis of this substrate are 0.347 mM and 1141 min(-1) respectively. The enzyme was affected by surface active agents to varying extents. The enzyme is stable for 2 h at 30 degrees C and pH 10.5. AP22 is also stable for 5 days over the pH range 9.0-11.0 at room temperature. AP22 has good pH stability compared with the alkaline proteases belonging to other strains of Bacillus clausii reported in the literature.

An alkaline protease from halotolerant Bacillus licheniformis BA17, isolated from Van Lake in Turkey, was purified 5.4 fold with 58% yield. The molecular weight was 19.7 kDa and the optimum temperature and pH were 60 °C and 10, respectively. The halflife of the pure enzyme was 38 h, 93 min, 14 min and 6 min at 40, 50, 60 and 70 °C, respectively. BA17 protease is very active at 30 °C between pH 8.0 and 10. Enzyme activity increased in the presence of Cu +2, Mg +2, Mn +2 and K +1 ions. Enzyme retained activity with 5% SDS (w/v) and 1% Triton X-100 (v/v). Inhibition with PMSF and EDTA suggested that the enzyme is a serine protease and is a metal-activated enzyme. Based on the N-terminal sequence of the first 13 amino acids, B. licheniformis BA17 alkaline protease did not show identity to any of those from other Bacillus species.

An extracellular serine alkaline protease of Bacillus clausii GMBAE 42 was produced in protein-rich medium in shake-flask cultures for 3 days at pH 10.5 and 37 degrees C. Highest alkaline protease activity was observed in the late stationary phase of cell cultivation. The enzyme was purified 16-fold from culture filtrate by DEAE-cellulose chromatography followed by (NH(4))(2)SO(4) precipitation, with a yield of 58%. SDS-PAGE analysis revealed the molecular weight of the enzyme to be 26.50 kDa. The optimum temperature for enzyme activity was 60 degrees C; however, it is shifted to 70 degrees C after addition of 5 mM Ca(2+) ions. The enzyme was stable between 30 and 40 degrees C for 2 h at pH 10.5; only 14% activity loss was observed at 50 degrees C. The optimal pH of the enzyme was 11.3. The enzyme was also stable in the pH 9.0--12.2 range for 24 h at 30 degrees C; however, activity losses of 38% and 76% were observed at pH values of 12.7 and 13.0, respectively. The activation energy of Hammarsten casein hydrolysis by the purified enzyme was 10.59 kcal mol(-1) (44.30 kJ mol(-1)). The enzyme was stable in the presence of the 1% (w/v) Tween-20, Tween-40,Tween-60, Tween-80, and 0.2% (w/v) SDS for 1 h at 30 degrees C and pH 10.5. Only 10% activity loss was observed with 1% sodium perborate under the same conditions. The enzyme was not inhibited by iodoacetate, ethylacetimidate, phenylglyoxal, iodoacetimidate, n-ethylmaleimidate, n-bromosuccinimide, diethylpyrocarbonate or n-ethyl-5-phenyl-iso-xazolium-3'-sulfonate. Its complete inhibition by phenylmethanesulfonylfluoride and relatively high k (cat) value for N-Suc-Ala-Ala-Pro-Phe-pNA hydrolysis indicates that the enzyme is a chymotrypsin-like serine protease. K (m) and k (cat) values were estimated at 0.655 microM N-Suc-Ala-Ala-Pro-Phe-pNA and 4.21 x 10(3) min(-1), respectively.

Properties of osmolytes that help protect biological molecules from stress and preserve their functions impose significant importance to finding new organisms with different osmolyte accumulation strategies. To this end, the moderately halophilic Halomonas sp. AAD12 has been characterized for its adaptation to stress conditions with particular emphasis on its osmolyte accumulation strategy. The effect of temperature, salinity, aeration and organic components on the accumulation of osmoprotectants and synthesis of fatty acids were examined in M63 minimal medium. Ectoine, proline and hydroxyectoine were the major osmolytes and palmitic acid (16:0), palmitoleic acid (16:1), and oleic acid (18:1) were the major fatty acids. Overall, ectoine yield was the highest among the three osmolytes at all salt concentrations and temperatures investigated. However, high salinity reduced ectoine yield with a concomitant increase in hydroxyectoine yield. The yield of hydroxyectoine as high as 525 mol/g dry cell mass at 37°C suggested that this microorganism could be a promising candidate as a hydroxyectoine producer.

In this study, our investigations showed that the increasing concentrations of all examined mono alcohols caused a decrease in the V m, k cat and k cat/K m values of Bacillus clausii GMBE 42 serine alkaline protease for casein hydrolysis. However, the K m value of the enzyme remained almost the same, which was an indicator of non-competitive inhibition. Whereas inhibition by methanol was partial non-competitive, inhibition by the rest of the alcohols tested was simple non-competitive. The inhibition constants (K I) were in the range of 1.32–3.10 M, and the order of the inhibitory effect was 1-propanol>2-propanol>methanol>ethanol. The ΔG ≠ and ΔG ≠ E − T values of the enzyme increased at increasing concentrations of all alcohols examined, but the ΔG ≠ ES value of the enzyme remained almost the same. The constant K m and ΔG ≠ ES values in the presence and absence of mono alcohols indicated the existence of different binding sites for mono alcohols and casein on enzyme the molecule. The k cat of the enzyme decreased linearly by increasing log P and decreasing dielectric constant (D) values, but the ΔG ≠ and ΔG ≠ E − T values of the enzyme increased by increasing log P and decreasing D values of the reaction medium containing mono alcohols.

Four Gram-negative, moderately halophilic, exopolysaccharide-producing strains, designated AAD6T, AAD4, AAD17 and AAD21, were isolated from Çamaltı Saltern Area, a wildlife reserve in Sasalı, İzmir province located in the Aegean Region of Turkey. The isolates grew at an optimum NaCl concentration of 10 % (w/v). The major cellular fatty acids were C16 : 0, C18 : 1ω7c, C16 : 1ω7c and C12 : 0 3OH, respectively and the predominant lipoquinone was ubiquinone Q-9. The G+C content of the genomic DNA of strains AAD6T, AAD4, AAD17 and AAD21 was 63.0, 63.3, 62.8 and 62.6 mol%, respectively. Comparative 16S rRNA gene sequence studies showed that the isolates belonged to the genus Halomonas . The DNA–DNA hybridization mean values between the representative strain AAD6T and the closely related species Halomonas salina DSM 5928T, Halomonas halophila DSM 4770T, Halomonas maura DSM 13445T, Halomonas organivorans DSM 16226T, Halomonas elongata DSM 2581T, Halomonas koreensis JCM 12237T and Halomonas nitroreducens LMG 24185, were 40.8, 39.6, 24.2, 23.3, 12.6, 14.5 and 12.2 %, respectively. Based on these data the strains represent a novel species of the genus Halomonas for which the name Halomonas smyrnensis sp. nov. is proposed. The type strain is AAD6T ( = DSM 21644T = JCM 15723T).

The moderately halophilic Halomonas sp. strain AAD21, which produces extracellular thermostable alpha-amylase, was isolated from the Camalti Saltern area located in Izmir, Turkey. NaCl, carbon, and nitrogen sources in the growth medium were optimized to enhance alpha-amylase yield. The highest enzyme yield was measured in the presence of 20% NaCl with peptone as the nitrogen and starch as the carbon sources in the fermentation broth. This microorganism was also found to utilize waste potato peel as a carbon source for alpha-amylase production. Concentrations of carbon and nitrogen sources were optimized using a statistical approach, and alpha-amylase activity increased from 4.07 U mL(-1) min(-1) to 26.25 U mL(-1) min(-1). Maximum alpha-amylase production was achieved at the end of 48 h of growth in the presence of 20% NaCl, 4.12% starch, 1.0% peptone, 0.2% KCl, 2% MgSO4 center dot 7H(2)O, and 0.03% trisodium citrate pentahydrate. The optimum pH and temperature of the alpha-amylase were found to be 7.0 and 50 degrees C, respectively. The alpha-amylase synthesized by Halomonas sp. AAD21 was also thermostable. Crude alpha-amylase did not lose its original activity after 2 h of incubation at 50 degrees C and 60 degrees C, and it retained 70% of its original activity after 120 min of incubation at 90 degrees C.

A gram-negative, moderately halophilic bacterium was isolated from Çamaltı Saltern area, located in the Aegean Region of Turkey. Analysis of its 16S rRNA gene sequence and physiological characteristics showed that this strain belonged to the genus Halomonas ; hence, it was designated as Halomonas sp. strain AAD12. The isolate tolerated up to 800 mg⋅L–1phenol; however, at elevated concentrations, phenol severely retarded cell growth. The increase in lag phase with increasing phenol concentrations indicated that the microorganism was undergoing serious adaptative changes. To understand the physiological responses of Halomonas sp. strain AAD12 to phenol, a 2-dimensional electrophoresis approach combined with mass spectrometric analysis was used. This approach showed that the expression of 14 protein spots were altered as phenol concentration increased from 200 to 800 mg⋅L–1. Among the identified proteins were those involved in protein biosynthesis, energy, transport, and stress metabolism. So far, this is the first study on phenolic adaptation of a gram-negative, moderately halophilic bacteria using proteomic tools. The results provided new insights for understanding the general mechanism used by moderately halophilic bacteria to tolerate phenol and suggested the potential for using these microorganisms in bioremediation.

A Gram-stain-positive, obligately alkaliphilic bacterium designated strain GMBE 72T was isolated from mushroom compost from Yalova, located in the Marmara region of Turkey. Cells were aerobic, straight rods and they formed subterminal to terminal ellipsoidal endospores. The isolate was catalase-positive, oxidase-negative and motile and contained a type A1γ peptidoglycan based on meso-diaminopimelic acid. The strain grew at pH 8.0–12.5. The major cellular fatty acid was anteiso-C15 : 0. The genomic DNA G+C content was 40.2 mol%. Phylogenetic analyses based on 16S rRNA gene sequencing showed that strain GMBE 72T belonged to the genus Bacillus and exhibited 98.2 % sequence similarity to Bacillus pseudofirmus DSM 8715T. DNA–DNA reassociation was 56 % between GMBE 72T and B. pseudofirmus DSM 8715T. According to our polyphasic characterization, strain GMBE 72T represents a novel species of the genus Bacillus, for which the name Bacillus marmarensis sp. nov. is proposed. The type strain is GMBE 72T (=DSM 21297T =JCM 15719T).

An alkaline protease producer Bacillus licheniformis strain was isolated from Van Lake in Turkey. The strain is Gram positive, aerobic, motile, sporulating rod-shaped bacterium. Spores were ellipsoidal and positioned central in nonswollen sporangium. The cells were able to grow well at a pH range of 5.7-10. The optimal growth temperature was found to be 37 °C. Growth at a wide range of NaCl concentration (from 0 to 20%) showed that BA17 is halotolerant. Main fatty acid composition of BA17 was anteiso-C15:0 and iso-C15:0. The strain was presumptively identified as B. licheniformis according to 16S rDNA gene sequence analysis. The most appropriate medium for the growth and protease production is composed of 0.5% yeast extract, 0.5% NaNO 3, 0.02% MgSO4·7H2O, 0.1% K 2HPO4 and 0.5% maltose. The optimum temperature and pH of the alkaline protease of strain BA17 were found to be 60 °C and pH 11, respectively. The activity was completely lost in the presence of PMSF, suggesting that the preparation contains serine-alkaline protease(s).

A . A . D E N I Z C I , D . K A Z A N , E . C . A . A B E L N A N D A . E R A R S L A N . 2003. Aims: The isolation and identification of new Bacillus sp. capable of growing under highly alkaline conditions as alkaline protease producers. Methods and Results: A Bacillus strain capable of growing under highly alkaline conditions was isolated from compost. The strain is a Gram-positive, spore-forming, motile, aerobic, catalase- and oxidase-positive, alkaliphilic bacterium and designated as GMBAE 42. Good growth of the strain was observed at pH 10. The strain was identified as Bacillus clausii according to the physiological properties, cellular fatty acid composition, G + C content of genomic DNA and 16S rRNA gene sequence analyses. The result of 16S rRNA sequence analyses placed this bacterium in a cluster with B. clausii. The G + C content of the genomic DNA of the isolate GMBAE 42 was found to be 49 mol%. The crude extracellular alkaline protease produced by the isolate showed maximal activity at pH 11AE0 and 60� C. Conclusions: The results suggest that isolated strain GMBAE 42 is a new type of B. clausii capable of growing at pH 10AE0 and produce extracellular alkaline protease very active at pH 11AE0. Significance and Impact of the Study: Isolated strain could be used in commercial alkaline protease production and its enzyme can be considered as a candidate as an additive for commercial detergents.