Isolation and identification of ß-galactosidase producing fungi and investigating the activity of enzyme from the selected isolate

ß-galactosidases are enzymes of the glycoside hydrolase family EC (3.2.1.23) that catalyze the hydrolysis of some disaccharides with wide applications in the food industry. Also, some ß-galactosidases have transgalactosylation activity. This study aimed to isolate ß-galactosidase-producing fungi from several traditional cheese and whey samples and to determine the enzymatic activity under the various conditions (temperature, time, pH, and different cations).

Yeasts and molds were isolated and screened for the ability to produce beta-galactosidase by a chromogenic test containing 5-bromo-4-chloro-3-indolyl-Beta-D-galacto-pyranoside (X-gal) in the media. After screening, the most promising isolated molds and yeasts were identified genotypically by ITS rDNA sequencing. Then, the hydrolytic activity of crude beta-galactosidase was evaluated using ortho-Nitrophenyl-β-galactoside (ONPG) as the substrate under different conditions (different levels of temperature, time, pH, and various cations Mn2+, Mg2+, Zn2+, Cu2+). Finally, the possibility of transgalactosylation activity of the selected enzyme was investigated.

The results of PCR product sequencing resulted in the identification of isolates MM24, MT12, and MW14 as Kluyveromyces lactis H1-3 (99.57%), Kluyveromyces lactis E3 (97.53%), and Penicillium brevicompactum (100%), respectively. Investigating the activity of ß-galactosidase produced by MM24, MT12, and MW14 isolates showed a higher activity for the crude enzyme from K. lactis MM24. Moreover, the highest activity was found at 37°C, pH 7, and 30 min reaction time. Different cations did not significantly influence the enzymatic activity at 0.1 and 1mM, however it was completely inhibited against Cu+2 and Zn+2 at 10 Mm. Furthermore, 10 mM Mn+2 and Ca+2 inhibited enzymatic activity by about 80% and 65%, respectively. High-performance liquid chromatography (HPLC) analysis confirmed ß-galactosidase activity on lactose, whereas no transgalactosylation activity was observed in this study.

Overall, K. lactis MM24 isolated in the current study can be considered as a new isolate for ß-galactosidase production.

ß-galactosidases are enzymes of the glycoside hydrolase family EC (3.2.1.23) that catalyze the hydrolysis of some disaccharides with wide applications in the food industry. Also, some ß-galactosidases have transgalactosylation activity. This study aimed to isolate ß-galactosidase-producing fungi from several traditional cheese and whey samples and to determine the enzymatic activity under the various conditions (temperature, time, pH, and different cations).

Yeasts and molds were isolated and screened for the ability to produce beta-galactosidase by a chromogenic test containing 5-bromo-4-chloro-3-indolyl-Beta-D-galacto-pyranoside (X-gal) in the media. After screening, the most promising isolated molds and yeasts were identified genotypically by ITS rDNA sequencing. Then, the hydrolytic activity of crude beta-galactosidase was evaluated using ortho-Nitrophenyl-β-galactoside (ONPG) as the substrate under different conditions (different levels of temperature, time, pH, and various cations Mn2+, Mg2+, Zn2+, Cu2+). Finally, the possibility of transgalactosylation activity of the selected enzyme was investigated.

The results of PCR product sequencing resulted in the identification of isolates MM24, MT12, and MW14 as Kluyveromyces lactis H1-3 (99.57%), Kluyveromyces lactis E3 (97.53%), and Penicillium brevicompactum (100%), respectively. Investigating the activity of ß-galactosidase produced by MM24, MT12, and MW14 isolates showed a higher activity for the crude enzyme from K. lactis MM24. Moreover, the highest activity was found at 37°C, pH 7, and 30 min reaction time. Different cations did not significantly influence the enzymatic activity at 0.1 and 1mM, however it was completely inhibited against Cu+2 and Zn+2 at 10 Mm. Furthermore, 10 mM Mn+2 and Ca+2 inhibited enzymatic activity by about 80% and 65%, respectively. High-performance liquid chromatography (HPLC) analysis confirmed ß-galactosidase activity on lactose, whereas no transgalactosylation activity was observed in this study.

Overall, K. lactis MM24 isolated in the current study can be considered as a new isolate for ß-galactosidase production.