Purification and characterization of a thermostable Galium aparine β-galactosidase: A competent agent with enhanced cytotoxic activity against MCF-7 cell line

• Published in: Process biochemistry (1991) 2024-12, Vol.147, p.39-50
• Main Authors: Alghamdi, Suad A., Rahman, Khalil ur, Zaman, Umber, Alissa, Mohammed, Al-Saeedi, Sameerah I., Alghamdi, Abdullah, Alghamdi, Amani, kelabi, Eman, Maashi, Marwah Suliman, Alsalmi, Ohud
• Format: Article
• Language: English
• Subjects: Gallium aparine; MCF-7 breast cancer cells; Therapeutic efficacy
• ISSN: 1359-5113
• DOI: 10.1016/j.procbio.2024.08.010

β-galactosidases (EC 3.2.1.23) are capable of catalysing two distinct types of reactions: transgalactosylation and hydrolysis. It is very important for the senescence of cancer cells. It is a lysosomal exoglycosidase involved in the catabolism of glycoconjugates by sequential release of β-linked terminal galactosyl residues. It has profound significance in cancer cell senescence. It can be derived from various sources including plants, animals, bacteria, yeasts, and filamentous fungi. Purification and thermodynamic characterization of β-galactosidase from Galium aparine seeds, and in vitro efficacy assessment of a breast cancer cell line. The enzyme was purified by using ammonium sulphate precipitation, dialysis, DEAE cellulose, and Sephadex G-100 chromatography. The enzyme was purified to 6.4 fold with a yield of 14 % and specific activity of 11.6 U/mg of protein. SDS-PAGE and MALDI-TOF analysis revealed that β-galactosidase is a monomer with a molecular weight of 35 kDa. The optimum pH and temperature for the purified enzyme were 6.0 and 50 ºC. The enzyme had Km and Vmax values of 0.25 mM and 32 µmol/min, respectively. The thermodynamic parameters (ΔHº, ΔGº, ΔSº) for the irreversible denaturation of the enzyme were also investigated at different temperatures (50–75 ºC). Lactose in milk was successfully hydrolyzed using a purified enzyme at a rate of 0.0121 min−1 and 12.12 min. This is better than other studied β-galactosidases. These results suggest the use of β-galactosidase from G. aparine for producing delactosed milk for the lactose-intolerant population and prebiotic synthesis. pH and optimum temperature and its activation by Ca2+ shows that it is suitable for milk processing. As a result, the enzyme was stable even at higher temperatures and had the potential to undergo catalysis, making it economically viable, primarily in the food and pharmaceutical industries. The present study is aimed to purification and thermodynamic characterization of enzyme to assess in vitro efficacy of β-galactosidase on MCF-7 cell line to delineate its therapeutic efficacy. [Display omitted] •β-galactosidase from prebiotic Galium aparine was purified 6.4 fold with 14 % yield.•β-galactosidase shows an indicator of the switch mechanism used by cells to enter senescence.•The purified enzyme hydrolyzed the milk lactose with a t1/2 of 14.74 min.•The treatment of the MCF-7 cell line with β-galactosidase inhibits cell growth•Thermostable industrially important enzyme from preb