Strain improvement of a low fructosyltransferase-producing Aureobasidium pullulans NAC8 (Accession No. KX023301) was carried out using chemical mutagens such as ethidium bromide and ethyl methane sulfonate. The wild-type and mutant strain were distinguished using Random amplified polymorphic DNA PCR and DNA fingerprinting analysis. Plackett-Burman and Box Behnken design were statistical tools used to determine important media parameters and optimization, respectively. Phenotypically and genetically, the new improved strain was different from the wild-type. The most important media parameters from PDB influencing fructosyltransferase production were ammonium chloride, sucrose and yeast extract at p < 0.05. Some significant parameters obtained with the BBD exhibited quadratic effects on FTase. The F values (35.37 and 32.11), correlation coefficient (0.98 and 0.97) and the percent coefficient of variation (2.53% and 2.40%) were obtained for extracellular and intracellular FTase respectively. The validation of the model in the improved strain resulted in an overall 6.0 and 2.0-fold increase in extracellular and intracellular FTase respectively compared to the wild-type.A relatively low FTase-producing strain of Aureobasidium pullulans NAC8 was enhanced for optimum production using a two-pronged approach involving mutagenesis and statistical optimization. The improved mutant strain also had remarkable biotechnological properties that make it a suitable alternative than the wild-type.

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In this study, the preparation, characterization, and evaluation of the cross-linked enzyme aggregates (CLEAs) of fructosyltransferase isolated from the mutant-type Aureobasidium pullulans NAC8 for the production of fructooligosaccharides (FOS) with prebiotic properties were investigated. The reaction conditions were optimized statistically by using response surface methodology. CLEAs were characterized by using both photo-microscopy and scanning electron microscopy energy-dispersive X-ray spectroscopy (SEM-EDX). The secondary structure of the protein was predicted by using Fourier transform infrared spectroscopy (FTIR). FOSs produced as a result of the biotransformation of sucrose was quantified by using a chemometric analysis of FTIR spectral data. The prepared FOSs were investigated for the prebiotic effects under anaerobic conditions using a standard strain of Lactobacillus sp. Following were the statistically determined, optimum conditions for the production of FOSs: 5% (v/v) glutaraldehyde, pH 5.5, and a temperature of 32EC. The values of R 2 , F value, adjusted precision, and coefficient of variation were found to be 0.94, 11.50, 9.34, and 35.34%, respectively. Validation of the model resulted in the reusability of the CLEAs in six reaction cycles as it retained 70% of the residual activity. The SEM-EDX and photomicrographic analysis showed that the sizes of CLEAs varied, which ranged between 10 and 40 μm. Aggregation leads to the formation of a higher number of beta sheets in the CLEAs than that in the free enzyme. The total FOSs produced by the CLEAs was around 57% (w/v). A. pullulans FOS supplemented to the medium had a stimulatory effect on the growth of bi dobacteria compared with the control medium. The growth curves of Lactobacillus sp. reached the plateau phase after 24 h of cultivation with FOS. The CLEA of fructosyltransferase catalyzed the biocatalysis of sucrose to FOSs, a process with a biotechnological importance as prebiotics.

Cellulase enzyme was purified and characterized from termite soldiers (Ametermes eveuncifer) using 70% ammonium sulphate precipitation, ion exchange chromatography and affinity chromatography. The enzyme isolated had a specific activity of 5.04 U/mg with a percentage yield of 11.7%. The enzyme showed maximum activity at 50 0 C and pH 8. The enzyme was not inhibited by Ba 2+ at a concentration of 1mM and Pb 2+ at 10 mM concentration but was inhibited by other metal ions at 1 mM and 10 mM concentrations of their salts (NaCl, KCl, MnCl 2 , and NiCl 2 ) ,

This study investigated the screening, production and optimization of an extracellular lipase from a fungus isolated from the contaminated soil of a palm oil processing shed. This was with a view to obtaining a strain that can secrete lipase with biochemical properties exploitable for biotechnological applications such as bioremediation of oil contaminated sites. Soil samples were collected from palm oil contaminated sites in Gbogan, Osun State, Nigeria (Latitude N 7°29.1481´ and Longitude E 4°20.7587´). The isolated fungal strains were screened on tributyrin agar for exogenous lipolytic activity. Molecular identification was carried out by amplifications of ITS-1, 5.8S and ITS-2 regions. The effects of incubation time, inducers, pH, temperature, carbon and nitrogen sources were varied for optimal lipase production using one factor at a time approach. Rhizopus oryzae ZAC3 (NCBI accession No: KX035094) was identified as the highest lipaseproducing strain. Maximum lipase production was observed on the fourth day, pH 5.0 and a temperature of 45 o C. Olive oil, xylose and yeast extract were the best inducer, carbon and nitrogen sources respectively for lipase production. There was a 2.02 fold increase in lipase production under these optimized conditions. In conclusion, Rhizopus oryzae ZAC3 lipase has properties exploitable for industrial and biotechnological applications.

Background β-Cyanoalanine synthase plays essential roles in germinating seeds, such as in cyanide homeostasis. Methods β-Cyanoalanine synthase was isolated from sorghum seeds, purified using chromatographic techniques and its biochemical and catalytic properties were determined. Results The purified enzyme had a yield of 61.74% and specific activity of 577.50 nmol H2S/min/mg of protein. The apparent and subunit molecular weight for purified β-cyanoalanine synthase were 58.26±2.41 kDa and 63.4 kDa, respectively. The kinetic parameters with sodium cyanide as substrate were 0.67±0.08 mM, 17.60±0.50 nmol H2S/mL/min, 2.97×10−1 s−1 and 4.43×102 M−1 s−1 for KM, Vmax, kcat and kcat/KM, respectively. With L-cysteine as substrate, the kinetic parameters were 2.64±0.37 mM, 63.41±4.04 nmol H2S/mL/min, 10.71×10−1 s−1 and 4.06×102 M−1 s−1 for KM, Vmax, kcat and kcat/KM, respectively. The optimum temperature and pH for activity were 35°C and 8.5, respectively. The enzyme retained more than half of its activity at 40°C. Inhibitors such as HgCl2, EDTA, glycine and iodoacetamide reduced enzyme activity. Conclusion The biochemical properties of β-cyanoalanine synthase in germinating sorghum seeds highlights its roles in maintaining cyanide homeostasis.

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