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Goa is situated in the northwestern part of the Western Dharwar Craton (WDC) which is Asia's major metallogenic province. The iron ore of Goa is associated with greenstone and occurs as bands, reefs, and lenses of Banded Hematite Quartzite (BHQ) and Banded Magnetite Quartzite (BMQ) none of these are explored for geo-microbiological dimensions, such as detection of microbioform Gold. The presence of metalliferous WDC in Goa affords sampling of auriferous materials from mining area having hitherto BHQ at latitude 15°29′54″ N and longitude 74° 03′44″ E) and BMQ at latitude 15°24′12″ N and longitude 74° 09′31″ E were collected, surface sterilized and drilled with a sterile driller to get the endolithic material under sterile conditions and sent for metagenomic analysis using Oxford nanopore sequencing. Both the samples showed the presence of microorganisms with Archea, such as Haloferax, Nitrososphaera, and Eubacteria, i.e., Bacillus, Ralstonia, Cupriavidus, Burkholderia, and Acetobacter. The traditional model of Banded Iron Deposition assumes precipitation by the oxidation of hydrothermal Fe (II), either via abiotic oxidation or biotic oxidation through chemo lithotrophic Eubacteria, such as cyanobacteria which produce oxygen. This paper reports the diversity and the presumptive role of these microorganisms in the biogeochemical cycling of metals, such as Fe, Mn, and gold (Au). [ABSTRACT FROM AUTHOR]

Industrial important proteases and lipases are in increasing demand for various biotechnological applications. In the present study, the concomitantly produced protease and lipase by Haloferax sp. strain GUBF 2 were simultaneously purified as a heterogeneous lipase (45 and 66 kDa) and homogeneous protease (180 kDa); with 28.3 and 31.36 fold purity, respectively using Sephadex G-200. The aforementioned extremozymes were active at pH 3–13, 20–80 °C, 1–5 M NaCl, with optimal activity at pH 6, 70 °C, and 3 M NaCl, thus exhibiting attributes of true haloextremozymes. The Km and Vmax of purified lipase were 3.47 mM and 16.2 U/mL, while protease were 3.29 mg/mL and 28.5 U/mL, respectively. FTIR bands corresponding to the vibrations of amide II and amide III were detected in haloextremozymes which could perhaps be used to determine the secondary structure of the purified proteins. Furthermore, the activity of both enzymes was stimulated by Ca2+ and inhibited by 10 mM Hg2+ and phenylmethyl sulphonyl fluoride (PMSF). Additionally, these haloextremozymes are stable in the presence of detergent additives and organic solvents. In addition, purified protease displayed 74.3 ± 4.85% in-vitro blood clot dissolution activity. Conclusively this study revealed the key features, unusual properties, and possible biomedical applications of detergent-stable and organic solvent-tolerant haloextremozymes from Haloferax sp. strain GUBF 2 to date unexplored. [ABSTRACT FROM AUTHOR]

Aim: To investigate the ability of Haloferax alexandrinus GUSF‐1 (KF796625) to biosynthesize non‐toxic elemental selenium (Se0) and check their capacity in in vitro crystal structure modulation of calcium oxalate, which are implicated in the development of renal calculi. Methods and Results: Haloferax alexandrinus GUSF‐1 (KF796625) during growth in the presence of 5 mmol L−1 of selenite formed insoluble brick‐red particles. Se0 formed was monitored spectrophotometrically using a combination of two assays; the ascorbic acid reduction and sodium sulphide solubilization assay. After 168 h of growth, 2.89 mmol L−1 of Se0 was formed from 4.9 mmol L−1 of selenite. Absorption bands at 1.5, 11.2 and 12.5 keV in EDX spectroscopy confirmed that the brick‐red particulate matter was Se0. Furthermore, these selenium nanoparticles (SeNPs) were pentagonal in shape in transmission electron microscopy imaging. The peak positions in X‐ray diffractogram at 2θ values of 23.40°, 29.66°, 41.26°, 43.68°, 45.24°, 51.62°, 55.93° and 61.47° and the relative intensities further confirmed the formation of Se0. In vitro addition of 50 and 100 µg ml−1 of these SeNPs to the mixture of sodium chloride, calcium chloride and sodium oxalate affected and modulated the shape and size of rectangular‐shaped calcium oxalate crystals (average area of 1.23 ± 0.2 µm2) to smaller rectangular‐shaped crystals (average area of 0.54 ± 0.2 µm2) and spherical‐shaped crystals (average area 0.13 ± 0.005 µm2). Conclusion: Haloferax alexandrinus GUSF‐1 (KF796625) transformed selenite to Se0 pentagonal nanoforms that modulated in vitro the formation of crystal shape and size of calcium oxalate. Significance and Impact of Study: There are no reports on conversion of selenite to Se0 among the Haloferax genera, and this study involving the formation of pentagonal SeNPs with capacity to modulate the formation of calcium oxalate crystals in haloarchaea is recorded as the first report and of significance in pharmaceutical research related to formulations abetting urinary calculi. [ABSTRACT FROM AUTHOR]

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%. Strain GUMF5 grew on mineral salts medium with 20% NaCl and 0.5% 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% cellulase with an activity of 131.13 U/mg and 1.24-fold purity was obtained. The purified cellulase had optimum activity at 20% NaCl, at 40 ºC, 0.5% 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+2 > Mn+2 > Mg+2 > SO42− > NH4+ and was inhibited by Ag+ > Fe+2 > Cu+2. Methanol and ethanol enhanced the cellulase activity by 6% and 26%, 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. [ABSTRACT FROM AUTHOR]

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%. Strain GUMF5 grew on mineral salts medium with 20% NaCl and 0.5% 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% cellulase with an activity of 131.13 U/mg and 1.24-fold purity was obtained. The purified cellulase had optimum activity at 20% NaCl, at 40 ºC, 0.5% 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+2 > Mn+2 > Mg+2 > SO42− > NH4+ and was inhibited by Ag+ > Fe+2 > Cu+2. Methanol and ethanol enhanced the cellulase activity by 6% and 26%, 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. [ABSTRACT FROM AUTHOR]

Hypersaline environment is a habitat with extreme osmotic conditions along with low Aw, serving as a home to extremely halophilic and halotolerant bacteria. The hypersaline environments, such as solar salterns located along the rivers, are exposed to fluxes of iron from iron ore transportation and other industrial wastes. The solar salterns often serve as a sink for metal intoxicants. Studies on archaea interaction with metal ions indicate the formation of minerals such as goethite, hematite, rhodochrosite, etc. However, studies exploring haloarchaeal candidates interacting with metals such as Fe3+ in a hypersaline growth condition are scarce. This study unveils for the first time formation of γFe2O3 from Fe3+ by the haloarchaeon, Haloferax sp. GUSF-1 thus implying the significance of the culture synthesizing minerals in hypersaline sediments. γFe2O3 is formed from Fe3+ by the haloarchaeon Haloferax sp. GUSF-1 (GenBank accession no.GU-1KF796625), under microaerophilic growth on sodium acetate. A 50 mg L−1 of Fe2+ and 30.6 mg L−1 of Fe3+ was detected inside the cells. Simultaneously, a brown-colored crystalline material deposited in the culture broth through an iron reductase inhibited by Zn2+ ions. The XRD of the deposit exhibited d values of 2.96, 2.514, 2.086, 1.6, and 1.45, while SEM-EDX displayed cubic and irregularly shaped minute particles with peaks for Fe at 0.6, 6.4, and 6.6 keV, respectively. TEM profiles revealed polycrystalline particles of 12–23 nm in size. Further, the SAED concentric pattern of light scattering with well-defined diffraction spots was consistent and matched with maghemite's crystal structure (γFe2O3). The FTIR spectrum revealed a peak at 1450 cm−1 indicating iron oxyhydroxide formation as an intermediate having γ-FeOOH stretching bond vibrations. Conclusively, this study opens the possibility of the haloarchaea isolated from solar salterns for its exploitation in nanobiotechnology. [ABSTRACT FROM AUTHOR]

• An extracellular inducible lipase and constitutive protease haloextremozymes were simultaneously produced by Haloferax lucentensis GUBF-2 MG076078. • The optimum NaCl for lipase haloextremozymes and protease production was 30 %, hence extremely halophilic. • Haloferax lucentensis GUBF-2 MG076078is a multienzyme producer and used for the valorization of agro-wastes. • Shrimp waste gave the highest protease, while coconut oil cake showed the highest lipase under optimal growth conditions. • The partially purified lipase and protease (called HLLPase) was used as Haloextremozyme-Bio-Additive in detergent formulation. Agro-food wastes rich in macronutrients are a replacement for expensive carbon and nitrogen sources used for enzyme production. In the present study, we aimed to valorize low-cost agro-wastes for the simultaneous production of protease and lipase haloextremozyme using Haloferax lucentensis GUBF-2 MG076078, characterization of agro-waste residues, and to apply haloextremozyme as Bio-additive in detergent formulation. It was observed that GUBF-2 efficiently hydrolyzed nine different agro-wastes with shrimp waste, and coconut oil cake yielded the highest protease (101.9 U/mL) and lipase (5.83 U/mL), respectively, under optimized conditions of 30 % NaCl, pH 6, and 42 °C. Further, SDS-PAGE of cell-free fermented agro-wastes revealed peptides from 212 to 34.6 kDa. Hydrolysates functioned as DPPH radical scavengers with maxima for shrimp waste (45.0 ± 1.15 %). SEM manifested structural alteration of fermented residues. FTIR showed degradation of proteins observed as 1640−1652 and 840−881 cm−1 absorptions with a fading peak at 1742 cm−1 indicating triacylglyceride hydrolysis. Furthermore, haloextremozyme-bio-additive detergent formulation efficiently removed 67.6 ± 3.19 % oil and 61.0 ± 3.60 % blood stains than 0.5 % Triton X-100. Conclusively, Haloferax lucentensis GUBF-2 MG076078 is a prospective strain for simultaneous protease and lipase haloextremozyme production from agro-food wastes, considering both cost efficiency and associated waste disposal problems. [ABSTRACT FROM AUTHOR]

Halococcus agarilyticus GUGFAWS-3 (MF425611) was isolated from a marine white sponge of Haliclona sp., inhabiting the rocks in the intertidal region of Anjuna, Goa, India. Uniquely, the microbe simultaneously produces two halo-extremozymes in 25% NaCl, namely protease and lipase at 49.5 ± 0.4 and 3.67 ± 0.02 (U mL−1), respectively. The protease is constitutively produced in starch mineral salts medium with consistent 4 ± 1.0 mm zone of enzyme production, regardless of the non-availability of protein as substrate. The ethanol precipitated enzyme on dialysis and Sephadex G-200 gel filtration chromatography was partially purified to 12.26-fold and was active between 20 and 80 °C, 0–5 M NaCl, and pH 3–13. Optimum activity, however, was at 70 °C, 3 M NaCl, and pH 7. The enzyme was thermo stable at 70 °C with 50.26 ± 2.40% of relative enzyme activity at 75 min. Furthermore, it was stable in the presence of polar and non-polar organic solvents, detergents, and hydrocarbons. Several metal cations enhanced its activity in the order of Ca2+ > Ni2+ > Fe3+ > Co2+ > Mg2+ > Cu2+ > Mn2+. Dependence of enzyme on cysteine; serine, and metal ions was confirmed by β-mercaptoethanol; PMSF and EDTA, respectively which induced its partial inhibition. Additionally, protease inhibited in vitro biofilm formation in Staphylococcus aureus. Conclusively, the production of a neutral halo-thermophilic protease is reported for the first time in the genus Halococcus. [ABSTRACT FROM AUTHOR]

Proteases and lipases are significant groups of enzymes for commercialization at the global level. Earlier, the industries depended on mesophilic proteases and lipases, which remain nonfunctional under extreme conditions. The discovery of extremophilic microorganisms, especially those belonging to haloarchaea, paved a new reserve of industrially competent extremozymes. Haloarchaea or halophilic archaea are polyextremophiles of domain Archaea that grow at high salinity, elevated temperature, pH range (pH 6–12), and low aw. Interestingly, haloarchaeal proteolytic and lipolytic enzymes also perform their catalytic function in the presence of 4–5 M NaCl in vivo and in vitro. Also, they are of great interest to study due to their capacity to function and are active at elevated temperatures, tolerance to pH extremes, and in non-aqueous media. In recent years, advances have been achieved in various aspects of genomic/molecular expression methods involving homologous and heterologous processes for the overproduction of these extremozymes and their characterization from haloarchaea. A few protease and lipase extremozymes have been successfully expressed in prokaryotic systems, especially E.coli, and enzyme modification techniques have improved the catalytic properties of the recombinant enzymes. Further, in-silico methods are currently applied to elucidate the structural and functional features of salt-stable protease and lipase in haloarchaea. In this review, the production and purification methods, catalytic and biochemical properties and biotechnological applications of haloextremozymes proteases and lipases are summarized along with recent advancements in overproduction and characterization of these enzymes, concluding with the directions for further in-depth research on proteases and lipases from haloarchaea. [ABSTRACT FROM AUTHOR]

Marine invertebrates exposed to high levels of reactive oxygen species in the oceans have been reported to produce antioxidants as a major defense against free radical mediated toxicity; protecting their tissues from the damage associated with the oxidative stress. In view of this, the present study was carried out to determine the antioxidant activity of 100 bacterial bionts isolated from marine sponges, corals and a single bivalve. Methanol extract of biont GUVFCFM-3 produced 67.83% scavenging of 2,2-diphenyl-2-picrylhydrazyl free radicals and 65.87% scavenging of superoxide free radicals. Preliminary tests leading to the identification of the extracellular antioxidant factor produced by GUVFCFM-3 revealed that it is a peptide. We report that the genera Chromohalobacter sp. primarily known for its unique salt tolerating abilities by virtue of the production of osmolytes is an excellent scavenger of free radicals. [ABSTRACT FROM AUTHOR]