Isolation of Halomicroarcula pellucida strain GUMF5, an archaeon from the Dead Sea-Israel possessing cellulase

Abstract

A strain designated GUMF5 was isolated in Goa-India from sediments of Dead Sea-Israel and identified as haloarchaeon Halomicroarcula pellucida based on 16S rRNA gene analysis similarity value of 99.84 percent. Strain GUMF5 grew on mineral salts medium with 20 percent NaCl and 0.5 percent carboxymethyl cellulose-sodium (CMC-Na) as a sole source of carbon and produced haloextremozyme cellulase. The enzyme was concentrated using Sephadex G20, precipitated with ethanol, dialyzed and retentate purified using Sephadex G200, the size exclusion chromatography. A yield of 78.53 percent cellulase with an activity of 131.13 U/mg and 1.24-fold purity was obtained. The purified cellulase had optimum activity at 20 percent NaCl, at 40 degrees C, 0.5 percent CMC-Na, pH 7 and 150 rpm. SDS-PAGE combined with zymographic analysis revealed the molecular weight of cellulase as 240 kDa, 40 kDa and 17.4 kDa. The activity of the enzyme was stimulated by metallic cations in the order of Ca sup(+2) greater than Mn sup(+2) greater than Mg sup(+2) greater than SO sub(4) sup(2-) grater than NH sub(4) sup(+) and was inhibited by Ag sup(+) greater than Fe sup(+2) greater than Cu sup(+2). Methanol and ethanol enhanced the cellulase activity by 6 percent and 26 percent, respectively. The haloextremozyme cellulase degraded Whatman No. 1 filter paper indicated in scanning electron micrographs, exposure of open pores and fibers without any intra connectivity corresponding to paperase activity and implicating the possible use of enzyme to bio-convert cellulosic waste. Conclusively, Halomicroarcula pellucida GUMF5 (Accession number: MH244431), globally, is the only Halomicroarcula pellucida isolated from the sediments of Dead Sea producing haloextremozyme cellulase, and hence is an important biotechnological resource.