Laccases couple the oxidation of phenolic compounds to the reduction of molecular oxygen and thus span a wide variety of applications. While laccases of eukaryotes and bacteria are well characterized, these enzymes have not been described in archaea. Here, we report the purification and characterization of a laccase (LccA) from the halophilic archaeon Haloferax volcanii. LccA was secreted at high levels into the culture supernatant of a recombinant H. volcanii strain, with peak activity (170 ؎ 10 mU ⅐ ml ؊1 ) at stationary phase (72 to 80 h). LccA was purified 13-fold to an overall yield of 72% and a specific activity of 29.4 U ⅐ mg ؊1 with an absorbance spectrum typical of blue multicopper oxidases. The mature LccA was processed to expose an N-terminal Ala after the removal of 31 amino acid residues and was glycosylated to 6.9% carbohydrate content. Purified LccA oxidized a variety of organic substrates, including bilirubin, syringaldazine (SGZ), 2,2,-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), and dimethoxyphenol (DMP), with DMP oxidation requiring the addition of CuSO 4 . Optimal oxidation of ABTS and SGZ was at 45°C and pH 6 and pH 8.4, respectively. The apparent K m values for SGZ, bilirubin, and ABTS were 35, 236, and 670 M, with corresponding k cat values of 22, 29, and 10 s ؊1 , respectively. The purified LccA was tolerant of high salt, mixed organosolvents, and high temperatures, with a half-life of inactivation at 50°C of 31.5 h.Multicopper oxidases (MCOs) are a family of enzymes that include laccases (p-diphenol: dioxygen oxidoreductases; EC 1.10.3.2), ascorbate oxidases (EC 1.10.3.3), ferroxidases (EC 1.16.3.1), bilirubin oxidases (EC 1.3.3.5), and other enzyme subfamilies (27,65). MCOs couple the oxidation of organic and/or inorganic substrates to the four-electron reduction of molecular oxygen to water. These enzymes often have four Cu atoms classified into type 1 (T1), type 2 (T2), and type 3 (T3) centers, in which a mononuclear T1 center on the surface of the enzyme provides long-range intramolecular one-electron transfer from electron-donating substrates to an internal trinuclear T2-T3 center formed by a T2 Cu coordinated with a T3 Cu pair. The T2-T3 cluster subsequently reduces dioxygen to water.Enzymes of the laccase subfamily oxidize a broad range of compounds, including phenols, polyphenols, aromatic amines, and nonphenolic substrates, by one-electron transfer to molecular oxygen and thus have a wide variety of applications from biofuels to human health. The best-known application is the use of a laccase from the lacquer tree Rhus vernicifera in paint and adhesives for more than 6,000 years in East Asia (29). Laccases have also been used in the delignification of pulp, bleaching of textiles and carcinogenic dyes, detoxification of water and soils, removal of phenolics from wines, improving adhesive properties of lignocellulosic products, determination of bilirubin levels in serum, and transformation of antibiotics and steroids (60). In addition, laccases have demonstrated pote...

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Strains of Halobacteria from an Algerian culture collection were screened for their lipolytic activity against p-nitrophenyl butyrate (PNPB) and p-nitrophenyl palmitate (PNPP). Most strains were active on both esters and 12% hydrolyzed olive oil. A strain identified as Natronococcus sp. was further studied. It grew optimally at 3.5 M NaCl, pH 8 and 40 degrees C. An increase in temperature shifted the optimum salt concentration range for growth from a wider range of 2-4 M, obtained at 25-30 degrees C, to a narrower range of 3.5-4 M, obtained at 35-40 degrees C. At 45 degrees C the optimum salt concentration was 2 M. These results show a clear correlation between salt and temperature requirement. The optimum conditions for the production of hydrolytic activity during growth were: 3.5 M NaCl and pH 8 for PNPB hydrolytic activity and 4 M NaCl and pH 7.5 for PNPP hydrolytic activity; both at 40 degrees C. The clear supernatant of cells grown at 4 M NaCl showed olive oil hydrolysis activity (in presence of 4 M NaCl) demonstrating the occurrence of a lipase activity in this strain. To our knowledge, this is the first report of a lipase activity at such high salt concentration.

Microbial populations within hypersaline lakes often exhibit high activities of photosynthesis, dissimilatory sulphate reduction and other processes and, thus, can have profound impacts on biogeochemical cycles of carbon, nitrogen, sulphur and other important elements within arid lands. To further understand these types of ecosystems, the physicochemical and biological properties of Sidi Ameur and Himalatt Salt Lakes in the Algerian Sahara were examined and compared. Both lakes were relatively neutral in pH (7.2 to 7.4) and high in salt, at 12% and 20 % (w/v) salinity for Himalatt and Sidi Ameur Lakes, respectively, with dominant ions of sodium and chloride. The community compositions of microbes from all three domains (Bacteria, Archaea and Eukarya) were surveyed through the use of 16S and 18S ribosomal gene amplification and clone library clustering using amplified ribosomal DNA restriction analysis (ARDRA) in conjunction with DNA sequencing and analysis. A high level of microbial diversity, particularly among the bacteria of the Himalatt Salt Lake and archaea of Sidi Ameur Lake, was found within these environments. Representatives from all known halophilic bacterial phyla as well as 6 different genera of halophilic archaea were identified. Moreover, several apparently novel phylotypes among both archaea and bacteria were revealed.

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Exopolysaccharides synthesized by lactic acid bacteria play a major role in the manufacturing of fermented dairy products as thickening agents. Exploration of the biodiversity of wild lactic acid bacteria from natural environments is currently the most suitable approach to search for the desired exopolysaccharide-phenotype. A total of 82 thermophilic lactic acid bacteria strains were isolated from Algerian raw camel milk. The isolation of strains was carried on modified Chalmers agar medium, under semi anaerobic conditions at 42°C. Bacterial isolates were phenotypically characterized and grouped into four genera: Lactobacillus (31.7%), Enterococcus (30.5%), Streptococcus (24.4%) and Pediococcus (13.4%). Based on the mucous type of the colonies, thirty EPS-positive strains were selected to be screened for their ability to produce exopolysaccharides. The production of polymers was carried out on Man, Rogosa and Sharpe (MRS) broth, supplemented with lactose and glucose as carbon sources. Yields quantification of soluble exopolysaccharides using a colorimetric method, showed that the selected strains produce yields ranging between 160 and 740 mg/l for Lactobacillus strains, between 126 and 319 mg/l for Streptococcus strains, between 70 and 242 mg/l for Enterococcus strains and between 132 and 134 mg/l for Pediococcus strains. This suggests that some strains have potential to be used as new culture starters for this and possibility other dairy products.

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