Insights into pH-induced conformational transition of β-galactosidase from Pisum sativum leading to its multimerization.
Although β-galactosidases are physiologically a very important enzyme and have may therapeutics applications, very little is known about the stability and the folding aspects of the enzyme. We have used β-galactosidase from Pisum sativum (PsBGAL) as model system to investigate stability, folding, and function relationship of β-galactosidases. PsBGAL is a vacuolar protein which has a tendency to multimerize at acidic pH with protein concentration ≥100 μg mL⁻¹ and dissociates into its subunits above neutral pH. It exhibits maximum activity as well as stability under acidic conditions. Further, it has different conformational orientations and core secondary structures at different pH. Substantial predominance of β-content and interfacial interactions through Trp residues play crucial role in pH-dependent multimerization of enzyme. Equilibrium unfolding of PsBGAL at acidic pH follows four-state model when monitored by changes in the secondary structure with two intermediates: one resembling to molten globule-like state while unfolding seen from activity and tertiary structure of PsBGAL fits to two-state model. Unfolding of PsBGAL at higher pH always follows two-state model. Furthermore, unfolding of PsBGAL reveals that it has at least two domains: α/β barrel containing catalytic site and the other is rich in β-content responsible for enzyme multimerization.
Dwevedi
Alka
A
School of Biotechnology, Faculty of Science, Banaras Hindu University, Varanasi 221005, India.
Dubey
Vikash Kumar
VK
Jagannadham
Medicherla V
MV
Kayastha
Arvind M
AM
D000818:Animals; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D057927:Hydrophobic and Hydrophilic Interactions; D007785:Lactose / Q000378:metabolism; D008892:Milk / Q000378:metabolism; D018532:Pisum sativum / Q000201:enzymology*; D010935:Plant Diseases; D017510:Protein Folding; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D058767:Protein Unfolding; D012639:Seeds / Q000201:enzymology; D001616:beta-Galactosidase / Q000737:chemistry* / Q000172:deficiency / Q000378:metabolism*
D001616:beta-Galactosidase; D007785:Lactose