Draft Genome Analysis of Acinetobacter indicus Strain UBT1, an Efficient Lipase and Biosurfactant Producer.

Acinetobacter indicus strain UBT1 has shown efficient lipase (243 U ml-1) and biosurfactant (61.1% E24% emulsification and surface tension reduction to 37.7 mN m-1) production capabilities using agro-industrial waste as sole carbon source. We report here the draft genome sequence of A. indicus strain UBT1 having genome size of 2.97 Mb with 45.90% GC content. Total 2721 coding genes were predicted using National Center for Biotechnology Information-Prokaryotic Genome Annotation Pipeline (NCBI-PGAP). The whole genome shotgun project sequence data are accessible through NCBI Gene Bank under accession no. JABFOI000000000. PGAP annotation revealed the presence of the triacylglycerol lipase, phospholipase etc., that circuitously confers the oil consumption competency to the strain UBT1. Rapid Annotation using the Subsystem Technology (RAST) server used for mapping the genes to the subsystem resulted in 278 subsystem with 30% subsystem coverage. The draft genome data can be used to exploit the A. indicus strain UBT1 for its advance biotechnological application and also for further comparative genomic studies.

Kinetic and thermodynamic characterization of a halotolerant ß-galactosidase produced by halotolerant Aspergillus tubingensis GR1.

ß-Galactosidase from halotolerant Aspergillus tubingensis GR1 was purified by two-step purification process comprising ammonium sulfate precipitation followed by size exclusion chromatography (SEC). The recovery of ß-galactosidase after SEC was found to be 1.40% with 58.55-fold increase in specific activity. The molecular weight of ß-galactosidase protein was found to be 93 kDa by SDS-PAGE. Activation energy for O-nitrophenol ß-D-galactopyranoside (ONPG) hydrolysis was 32.88 kJ mol(-1), while temperature quotient (Q(10)) was found to be 1.375. The enzyme was found to be stable over wide pH range and thermally stable at 60-65°C up to 60 min of incubation while exhibited maximum activity at 65°C with pH 3.0. V(max), K(m), and K(cat) for ONPG were found to be 2000 U ml(-1), 8.33 mM (ONPG), and 101454 s(-1), respectively. Activation energy for irreversible inactivation Ea(d) of ß-galactosidase was 100.017 kJ mol(-1). Thermodynamic parameters of irreversible inactivation of ß-galactosidase and ONPG hydrolysis were also determined. However, ß-galactosidase enzyme activity was activated significantly in the presence of 15% NaCl and hence shows activity up to 30% NaCl concentration.

Utilization of agro-industrial waste for ß-galactosidase production under solid state fermentation using halotolerant Aspergillus tubingensis GR1 isolate.

A halotolerant fungal isolate Aspergillus tubingensis GR1 was isolated from the man-made solar saltern located at Khambhat, Gujarat, India, and identified using 28S rDNA partial genome sequencing. This isolate was studied for ß-galactosidase production under solid state fermentation using wheat bran and deproteinized acid cheese whey. The influence of various agro-industrial wastes, nitrogen source and other growth conditions on ß-galactosidase production was investigated using 'one-factor-at-a-time' approach. Among various variables screened along with wheat bran and deproteinized acid cheese whey as major growth substrate, corn steep liquor and MgSO4 were found to be most significant. The optimum concentrations of these significant parameters were determined employing the response surface central composite design, revealing corn steep liquor concentration (2 mL) and magnesium sulphate (50 mg) per 5 g of wheat bran and 20 mL of deproteinized acid cheese whey for highest enzyme production (15,936 U/gds). These results suggest the feasibility of industrial large-scale production of ß-galactosidase known to be valuable in whey hydrolysis and removal of galactosyl residue from polysaccharide.

A statistical approach for the production of thermostable and alklophilic alpha-amylase from Bacillus amyloliquefaciens KCP2 under solid-state fermentation.

The bacterial strain producing thermostable, alklophilic alpha-amylase was identified as Bacillus amyloliquefaciens KCP2 using 16S rDNA gene sequencing data (NCBI Accession No: KF112071). Medium components were optimized through the statistical approach for the synthesis of alpha-amylase by the organism under solid-state fermentation using wheat bran as the substrate. The medium components influencing the enzyme production were identified using a two-level fractional factorial Plackett-Burman design. Among the various variables screened, starch, ammonium sulphate and calcium chloride were found to be most significant medium components. The optimum levels of these significant parameters were determined employing the response surface Central Composite design which significantly increased the enzyme production with the supplementation of starch 0.01 g, ammonium sulphate 0.2 g and 5 mM calcium chloride in the production medium. Temperature and pH stability of the alpha-amylase suggested its wide application in the food and pharmaceutical industries.

Kinetic and Thermodynamic Characterization of Glucoamylase from Colletotrichum sp. KCP1.

Extracellular glucoamylase of Colletotrichum sp. KCP1 produced through solid state fermentation was purified by two steps purification process comprising ammonium sulphate precipitation followed by gel permeation chromatography (GPC). The Recovery of glucoamylase after GPC was 50.40 % with 19.3-fold increase in specific activity. The molecular weight of enzyme was found to be 162.18 kDa by native-PAGE and was dimeric protein of two sub-units with molecular weight of 94.62 and 67.60 kDa as determined by SDS-PAGE. Activation energy for starch hydrolysis was 26.45 kJ mol(-1) while temperature quotient (Q 10 ) was found to be 1.9. The enzyme was found to be stable over wide pH range and thermally stable at 40-50 °C up to 120 min while exhibited maximum activity at 50 °C with pH 5.0. The pKa1 and pKa2 of ionisable groups of active site controlling V max were 3.5 and 6.8, respectively. V max , K m and K cat for starch hydrolysis were found to be 58.82 U ml(-1), 1.17 mg (starch) ml(-1) and 449 s(-1), respectively. Activation energy for irreversible inactivation (E a(d)) of glucoamylase was 74.85 kJ mol(-1). Thermodynamic parameters of irreversible inactivation of glucoamylase and starch hydrolysis were also determined.