Harvinder Singh Saini

Purification and characterization of β-glucosidase from Melanocarpus sp. MTCC 3922

This study reports the purification and characterization of β-glucosidase from a newly isolated thermophilic fungus, Melanocarpus sp. Microbial Type Culture Collection (MTCC) 3922. The molecular weight of β-glucosidase was determined to be ~ 92 and 102 kDa with SDS PAGE and gel filtration, respectively, and p I of ~ 4.1. It was optimally active at 60oC and pH 6.0, though was stable at 50oC and pH 5.0 - 6.0. The presence of DTT, mercaptoethanol and metal ions such as Na + , K + , Ca 2+ , Mg 2+ and Zn 2+ positively influenced the activity of β-glucosidase but the activity was inhibited in the presence of CuSO 4 . β-glucosidase recognized pNP- β-glucopyranoside (pNPG) as the preferred substrate, and showed very low affinity for pNP- β-D-cellobioside. K m and V max for the hydrolysis of pNPG by β-glucosidase was calculated as 3.3 mM and 43.68 µmolmin -1 mg protein -1 , respectively and k cat was quantified as 4 x 10 3 min -1 . β-glucosidase activity was enhanced appreciably in the presence of alcohols (methanol and ethanol) moreover, purified β-glucosidase showed putative transglycosylation activity that was positively catalyzed in presence of methanol as an acceptor molecule.

Isolation of hexachlorocyclohexane‐degrading Sphingomonas sp. by dehalogenase assay and characterization of genes involved in γ‐HCH degradation

Aim:  To screen and identify bacteria from contaminated soil samples which can degrade hexachlorocyclohexane (HCH)‐isomers based on dechlorinase enzyme activity and characterize genes and metabolites.

Biosurfactant production by Pseudomonas sp. and its role in aqueous phase partitioning and biodegradation of chlorpyrifos.

Aim:  To study the effect of biosurfactant on aqueous phase solubility and biodegradation of chlorpyrifos.

Efficient purification of the biosurfactant viscosin from Pseudomonas libanensis strain M9-3 and its physicochemical and biological properties.

Viscosin (1), an effective surface-active cyclic lipopeptide, was efficiently recovered from Pseudomonas libanensis M9-3 with a simple purification protocol. A major pigment also obtained during this process was identified as phenazine-1-carboxylic acid. The critical micelle concentration (cmc) of viscosin was determined to be 54 mg L (-1), and the minimum surface tension between air and water at the cmc was 28 mN m (-1). Viscosin forms stable emulsions even at low concentrations (7.5 mg L (-1)), and the conditional stability constant for a cadmium-viscosin complex was determined to be 5.87. The physicochemical properties measured for viscosin are similar to other well-studied biosurfactants such as rhamnolipid and surfactin. Viscosin inhibited migration of the metastatic prostate cancer cell line, PC-3M, without visible toxicity. These properties suggest the potential of viscosin in environmental and biomedical applications.

Molecular characterization of multiple xylanase producing thermophilic/thermotolerant fungi isolated from composting materials

Aims:  Molecular characterization of commercially important group of xylanase producing thermophilic/thermotolerant fungi.

Metabolic profiles and phylogenetic diversity of microbial communities from chlorinated pesticides contaminated sites of different geographical habitats of India.

Aims:  To study the microbial communities in three sites contaminated with chlorinated pesticides and evaluation of dehydrodechlorinase (linA) gene variants involved in gamma‐hexachlorocyclohexane (γ‐HCH, lindane) degradation.

Purification and characterization of two thermostable xylanases from Malbranchea flava active under alkaline conditions.

Two xylanases, MFX I and MFX II, from the thermophilic fungus Malbranchea flava MTCC 4889 with molecular masses of 25.2 and 30kDa and pIs of 4.5 and 3.7, respectively were purified to homogeneity. The xylanases were optimally active at pH 9.0 and at 60 degrees C, exhibited a half-life of 4h at 60 degrees C, and showed distinct mode of action and product profiles when applied to birchwood, oat spelt, and larchwood xylan, and to wheat and rye arabinoxylan. The xylanases were most active on larchwood xylan with K(m) values of 1.25 and 3.7mg/ml. K(cat)/K(m) values suggested that the xylanases preferentially hydrolyzed rye arabinoxylan. LC-MS/MS (liquid chromatography/mass spectrometry) analysis of tryptic digests of MFX I and MFX II revealed similarity with known fungal xylanases and suggests that that they belonged to the GH 11 and 10 glycosyl hydrolase super families, respectively. These xylanases can potentially be used in enzyme-assisted bleaching of the pulp derived from agro-residues, as well as production of xylooligosaccharides for pre-biotic functional food applications.

Amylase hyperproduction by deregulated mutants of the thermophilic fungus Thermomyces lanuginosus

Thermomyces lanuginosus was subjected to three cycles of mutagenesis (UV/NTG) and a selection procedure to develop amylase-hyperproducing, catabolite-repression-resistant and partially constitutive strains. One of the selected derepressed mutant strain III51, produced ∼7- and 3-fold higher specific activity of α-amylase (190 U/mg protein) and glucoamylase (105 U/mg protein), respectively, compared to a wild-type parental strain. Further, the effect of production parameters on mutant strain III51 was studied using a Box–Behnken design. The regression models computed showed significantly high R2 values of 96 and 97% for α-amylase and glucoamylase activities, respectively, indicating that they are appropriate for predicting relationships between corn flour, soybean meal and pH with α-amylase and glucoamylase production. Journal of Industrial Microbiology & Biotechnology (2002) 29, 70–74 doi:10.1038/sj.jim.7000270

Biological treatment of textile dye Acid violet‐17 by bacterial consortium in an up‐flow immobilized cell bioreactor

Aims:  To develop a cost effective and efficient biological treatment process for small scale textile processing industries (TPI) releasing untreated effluents containing intense coloured Acid violet‐17 (AV‐17), a triphenyl methane (TPM) group textile dye.

Xylanase production by Thermomyces lanuginosus wild and mutant strains

Thermomyces lanuginosus strains from different culture collections, namely ATCC 26909, ATCC 22083, DEN 1457, IMI 84400 and BS1 were compared for xylanase production, and isozyme profile. Of all the strains of T. lanuginosus, BS1 a soil isolate produced the largest amount of xylanase. All strains were found to produce two forms of xylanase (I & II) with molecular mass corresponding to 25.0 and 54.0 KDa. The u.v/NTG mutagenesis of T. lanuginosus BS1 aleurospores/protoplasts resulted in xylanase-hyperproducing mutants. A morphological colour mutant RB 524 produced approximately 2.5-fold higher xylanase (2506.0 units/ml) as compared to the parent strain (1018.1 units/ml).