Optimization of mannanase production from recombinant aspergillus sojae and analysis of galactomannan hydrolysis

Download

index.pdf

Date

2008

Author

Öztürk, Bengü

Metadata

Show full item record

Item Usage Stats

234
views

138
downloads

Aspergillus fumigatus produces enzymes required for the hydrolysis of galactomannans like locust bean gum. Among these enzymes endo-beta-1,4 mannanase is also produced at high levels. However, the fungus is not safe for use in the food industry. Therefore, the gene encoding endo-beta-1,4-mannanase of A. fumigatus IMI 385708 was previously cloned in our laboratory into Aspergillus sojae ATCC11906 which is a safe microorganism for use in food applications. Altogether eight transformants were obtained. It was shown that some of these transformants overproduce the enzyme because of expression under the control of glyceraldehyde-3-phosphate dehydrogenase promoter and fusion to the glucoamylase signal and pro-peptide coding region of Aspergillus niger. In this study, mannanase production of these transformants was compared with A. fumigatus and A. sojae transformant AsT1 showed c. 12 fold increase with the maximum activity of 352 U/ml. The effects of initial medium pH and number of spores on activity were investigated and maximum activity was achieved at pH 7.0 and the number of spores was found as 3.6 × 106. Optimization of the growth conditions for maximum mannanase production in shake flasks by using the best mannanase producing transformant AsT1 was carried out by using Box-Behnken design under Response Surface Methodology. The highest beta-mannanase activity on the fourth day of cultivation at 30 ºC was obtained as 363 U/ml in the optimized medium containing 7% sugar beet molasses, 0.43% NH4NO3, 0.1% K2HPO4, 0.05% MgSO4 as the weight/volume percentage at 207 rpm. On sixth day of cultivation under the optimized conditions, the highest mannanase activity was achieved as 482 U/ml which is 1.4 fold of 352 U/ml activity found on glucose medium previously. After 48 h of LBG hydrolysis by 40 U of mannanase, mannotriose, 61-galactosyl-beta-D-mannotriose and 63,64-di-alpha-galactosyl-beta-1,4-mannopentaose were found as the main products via HPLC analysis.

Subject Keywords

Food engineering., Recombinant Aspergillus Sojae.

URI

http://etd.lib.metu.edu.tr/upload/3/12609469/index.pdf
https://hdl.handle.net/11511/17851

Collections

Graduate School of Natural and Applied Sciences, Thesis

Suggestions

OpenMETU
Core

PREPARATION AND CHARACTERIZATION OF POLYMER COATED SMALL UNILAMELLAR VESICLES OZDEN, MY; Hasırcı, Vasıf Nejat (1991-09-02) Glucose oxidase was entrapped in small unilamellar vesicles composed of phosphatidylcholine, dicetyl phosphate and cholesterol. Prediction of the enzyme content of liposomes by calculations based on input concentrations of lipid and protein, dimensions of the lipids and the liposomes yielded one protein per vesicle. The entrapment efficiency was experimentally determined to be about 13%. On the other hand the entrapment efficiency for the small chromate ions was found to be significantly lower (0.1%). T...
Utilization of pretreated molasses for serine alkaline protease production with recombinant Bacillus species Calik, G; Pehlivan, N; Kalender, N; Ozdamar, TH; Çalık, Pınar (Informa UK Limited, 2003-05-01) Recombinant Bacillus amyloliquefaciens, Bacillus cereus, Bacillus subtilis , and Bacillus licheniformis were used for the production of serine alkaline protease (SAP) utilizing chemically and/or physically pretreated molasses. The highest enzyme activity was obtained with r- Bacillus subtilis , with the complex medium involving physically treated molasses having 20 kg m(-3) initial sucrose concentration in small-scale, agitation- and heating rate-controlled bioreactors at t=63 h. Effects of oxygen transfer ...
Monitoring the effects of divalent ions (Mn+2 and Ca+2) in heat-set whey protein gels Öztop, Halil Mecit; MCCARTHY, Michael J.; ROSENBERG, Moshe (Elsevier BV, 2014-04-01) Exploring the effects of cations in whey protein-based gels (WPG) is of importance when these gels are used for controlled release applications in food systems. The objective of this study was to evaluate both water uptake and cation release from heat-set WPGs. Magnetic Resonance Imaging and NMR relaxometry were employed to study the uptake and release. A non-paramagnetic (Ca+2) and a paramagnetic cation (Mn+2) were incorporated into the WPG as model divalent cations. Cylindrical pieces of WPGs with mangane...
Evaluation of high hydrostatic pressure sensitivity of Staphylococcus aureus and Escherichia coli O157 : H7 by differential scanning calorimetry Alpas, Hami; BOZOGLU, F; KALETUNC, G (Elsevier BV, 2003-11-01) Differential scanning calorimetry (DSC) was used to evaluate the relative high hydrostatic pressure (HHP) resistances of bacterial strains from Staphylococcus aureus and Escherichia coli O157:H7 in vivo. The total apparent enthalpy change and thermal stability were two DSC parameters used to compare bacterial strains of untreated control and pressure-treated bacteria. DSC thermograms indicated that ribosomal denaturation appears to be a major factor in cell death by both thermal and high pressure treatments...
Optimization studies on the features of an activated charcoal-supported urease system Kibarer, GD; Akovali, G (Elsevier BV, 1996-08-01) The adsorption of urease onto a well-defined solid support, petroleum-based activated charcoal, has been achieved to provide the enzymatic hydrolysis of urea. In order to produce a biocompatible surface, the enzyme support system has been coated with hexamethyldisiloxane through plasma polymerization. The quality of the resulting coat was tested by electronic spectroscopy for chemical analysis and scanning electron microscopy techniques. Studies on the adsorption of urease, and activity and stability of the...

Citation Formats

B. Öztürk, “Optimization of mannanase production from recombinant aspergillus sojae and analysis of galactomannan hydrolysis,” M.S. - Master of Science, Middle East Technical University, 2008.