Diversity of culturable moderately halophilic bacteria producing extracellular hydrolytic enzymes from marine sediments / Diversidade de bactérias cultiváveis moderadamente halofílicas que produzem enzimas hidrolíticas extracelulares de sedimentos marinhos

A total of 114 moderately halophilic bacteria were isolated from marine sediment environments. The isolates are belonged to 23 species based on the 16S rRNA sequence analysis. 63, 52, 47, 57, 74, 15 and 4 isolates are able to produce protease, amylase, lipase, pectinase, pulluanase, xylanase, cellulase, respectively. Combined hydrolytic enzyme activity analysis show that 15 strains present 1 hydrolytic activity, 32 strains present 2 hydrolytic activities, 21 strains present 3 hydrolytic activities, 26 strains present 4 hydrolytic activities, 11 strains present 5 hydrolytic activities and 2 strains present 6 hydrolytic activities. Hydrolase activities are widely distributed in a variety of species. The highest rates for production of protease, amylase, lipase, pectinase, pullanase, xylanase and cellulase were observed in species of B. baekryungensis, Hallobacillus sp., B. pumilus, B. megaterium or P. chungwhensis, B. amyloliquefaciens, B. pumilus, B. baekryungensis, respectively. However, the higher activities of protease, pectinase and pulluanase are frequently produced by the species of Halomonas sp. B. amyloliquefaciens or P. chungwhensis, and Vibrio sp. respectively. This investigation show that the diversity of halophilic bacteria from marine sediments could serve as a potential source of hydrolytic enzymes for industrial applications. (AU)

Um total de 114 bactérias moderadamente halofílicas foram isoladas de ambientes de sedimentos marinhos. Os isolados pertencem a 23 espécies com base na análise da sequência 16S rRNA. 63, 52, 47, 57, 74, 15 e 4 isolados são capazes de produzir protease, amilase, lipase, pectinase, pululanase, xilanase, celulase, respectivamente. A análise da atividade enzimática hidrolítica combinada mostra que 15 cepas apresentam 1 atividade hidrolítica, 32 cepas apresentam 2 atividades hidrolíticas, 21 cepas apresentam 3 atividades hidrolíticas, 26 cepas apresentam 4 atividades hidrolíticas, 11 cepas apresentam 5 atividades hidrolíticas e 2 cepas apresentam 6 atividades hidrolíticas. Atividades de hidrolase são amplamente distribuídas em uma variedade de espécies. As maiores taxas de produção de protease, amilase, lipase, pectinase, pululanase, xilanase e celulase foram observadas em espécies de B. baekryungensis, Hallobacillus sp., B. pumilus, B. megaterium ou P. chungwhensis, B. amyloliquefaciens, B. pumilus, B. baekryungensis, respectivamente. No entanto, as atividades mais elevadas de protease, pectinase e pululanase são freqüentemente produzidas pelas espécies de Halomonas sp. B. amyloliquefaciens ou P. chungwhensis e Vibrio sp. respectivamente. Esta investigação mostra que a diversidade de bactérias halofílicas de sedimentos marinhos pode servir como uma fonte potencial de enzimas hidrolíticas para aplicações industriais. (AU)

Proteolytic activity of recombinant DegP from Chromohalobacter salexigens BKL5

Background: DegP is a serine protease that specifically cleaves and refolds unfolding proteins in the periplasmic space of the cells. To date, there is no information regarding DegP from halophilic bacteria. Chromohalobacter salexigens BKL5 is a moderately halophilic bacterium that has the ability to grow in a media containing more than 15% salt. Therefore, the objectives of this work were to clone and overexpress DegP-encoding gene from C. salexigens BKL5 and characterize its biochemical properties. Results: DegP-encoding gene was overexpressed in Escherichia coli BL21(DE3) CodonPlus in an active form. SDS-PAGE analysis showed that the molecular weight of the recombinant DegP was 45 kDa. Size-exclusion chromatography analysis suggested that recombinant DegP was present in two multimeric states, hexameric and dodecameric, with molecular weights of 297.9 and 579.12 kDa, respectively. Both conformations were enzymatically active when casein was used as substrate for enzymatic assay. Circular dichroism analysis showed that recombinant DegP was composed of 0.21­0.29 helical content, which was comparable to the helical content in the crystal structure of E. coli DegP. The basic/acidic residue ratio of recombinant DegP was 0.56, which was slightly higher than that of DegP from extreme halophiles (average, 0.45) but significantly lower than that of DegP from nonhalophiles (average, 0.94). Conclusions: Recombinant DegP from C. salexigens BKL5 showed proteolytic activity when ß-casein was used as a substrate. In silico analysis indicated that recombinant DegP had characteristics similar to those of halophilic proteins depending on its amino acid composition.

NUMERICAL TAXONOMY OF MODERATELY HALOPHILIC BACTERIA ISOLATED FROM SALT LAKES IN XINJIANG REGION / 微生物学通报

Twenty nine moderately halophilic bacteria isolated from salt lakes in Xinjiang Region were compared with nine reference strains of relative species by performing a numerical analysis of 112 phenotypic characteristics. All the new isolates are Gram negative and can grow on the media containing 0～25% NaCl. The results showed that all tested strains were clustered into 2 phena at 76 % similarity level. Phenon I was composed by 25 new isolates strains,and phenon II included 9 reference strains and 4 new lsolated strains. Most of the new isolated strains were clusted into 3 subphena at 86 % similarity level. There were 10 new isolated strains in subphenon 1, 8 new isolated strains in subphenon 2, and 1 new isolated strain and 5 reference strains were included in subphenon 3. The results indicated that there were great differences between the new isolates and the reference strains.

Osmoregulation of a Halophilic Bacteria Strain Halomonas sp.BYS-1 / 微生物学通报

Halomonas sp.BYS-1 was a moderately halophilic bacteria strain isolated from activated sludge,It could grow on MM with NaCl concentration from 0.1～2.6 mol/L and phenyl acetic acid as sole carbon souce. When BYS-1 grew in the media with different concentrations of NaCl, there was no obvious change in its intracellular Na+ contents , it accumulated K+, glutamic acid and betaine as osmoprotectants. Its intracellular contents of K+,glutamic acid and betaine increased by 1.9,2.4 and 13.6 times, respectively, when the concentration of NaCl increased from 0.1 mol/L to 2mol/L.