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Applied Biochemistry and Biotechnology

, Volume 162, Issue 8, pp 2294–2312 | Cite as

Insights into pH-Induced Conformational Transition of β-Galactosidase from Pisum sativum Leading to its Multimerization

  • Alka Dwevedi
  • Vikash Kumar Dubey
  • Medicherla V. Jagannadham
  • Arvind M. KayasthaEmail author
Article

Abstract

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.

Keywords

PsBGAL Secondary structure Intrinsic fluorescence Hydrophobic domains 

Notes

Acknowledgement

A.D. would like to thank the Council of Scientific and Industrial Research (CSIR), New Delhi for financial assistance in the form of research fellowship.

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Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Alka Dwevedi
    • 1
  • Vikash Kumar Dubey
    • 2
  • Medicherla V. Jagannadham
    • 3
  • Arvind M. Kayastha
    • 1
    Email author
  1. 1.School of Biotechnology, Faculty of ScienceBanaras Hindu UniversityVaranasiIndia
  2. 2.Department of BiotechnologyIndian Institute of Technology GuwahatiGuwahatiIndia
  3. 3.Molecular Biology Unit, Institute of Medical SciencesBanaras Hindu UniversityVaranasiIndia

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