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Jack bean urease (EC 3.5.1.5) aggregation monitored by dynamic and static light scattering.
Cristian Follmer1, Fabiano V Pereira, Nádya P Da Silveira
1Department of Biophysics, IB, and Graduate Program in Cellular and Molecular Biology, Center of Biotechnology, Universidade Federal do Rio Grande do Sul, Av. Bento Gonalves 9500, Porto Alegre CEP 91501970, RS, Brazil.
Biophysical Chemistry
|September 29, 2004
Summary
Jack bean urease (JBU) aggregation alters its biological activities. Freeze-thawing causes irreversible JBU aggregation into high-order oligomers, unaffected by reducing agents or salts.
Area of Science:
- Biochemistry
- Enzymology
- Protein Chemistry
Background:
- Jack bean urease (JBU) aggregation impacts its biological functions, including ureolytic and entomotoxic activities.
- Understanding JBU aggregation is crucial for elucidating its diverse biological roles.
Purpose of the Study:
- To characterize protein aggregates of JBU using light scattering techniques.
- To investigate the influence of disulfide reducing agents and freeze-thawing cycles on JBU aggregation.
Main Methods:
- Dynamic Light Scattering (DLS) and Static Light Scattering (SLS) were employed.
- Determination of apparent hydrodynamic radii, average molecular masses, radii of gyration, and second virial coefficients.
- Assessment of the effects of dithiothreitol (DTT) and salt concentrations on JBU aggregation.
Main Results:
- Disulfide reducing agents did not affect JBU protein association, contrary to prior reports.
- Freeze-thawing cycles induced aggregation of native JBU (7 nm hydrodynamic radius, 12 nm radius of gyration) into high-order oligomers.
- The observed aggregation was irreversible by dithiothreitol (DTT) treatment or high salt concentrations.
Conclusions:
- JBU aggregation is a complex phenomenon influenced by environmental factors like freeze-thawing.
- The irreversibility of JBU aggregation under tested conditions provides insights into its solution behavior.
- Findings may correlate with the varied biological properties observed for JBU.