Thermal and Sodium Dodecylsulfate Induced Transitions of Streptavidin

Abstract

The strong specific binding of streptavidin (SA) to biotin is utilized in numerous biotechnological applications. The SA tetramer is also known to exhibit significant stability, even in the presence of sodium dodecylsulfate (SDS). Despite its importance, relatively little is known about the nature of the thermal denaturation pathway for SA. This work uses a homogeneous SA preparation to expand on the data of previous literature reports, leading to the proposal of a model for temperature induced structural changes in SA. Temperature dependent data were obtained by SDS and native polyacrylamide gel electrophoresis (PAGE), differential scanning calorimetry (DSC), and fluorescence and ultraviolet (UV)-visible spectroscopy in the presence and absence of SDS. In addition to the development of this model, it is found that the major thermal transition of SA in 1% SDS is reversible. Finally, although SA exhibits significant precipitation at elevated temperatures in aqueous solution, inclusion of SDS acts to prevent SA aggregation.

Engineering Soluble Monomeric Streptavidin with Reversible Biotin Binding Capability

“…First, streptavidin has stronger subunit interfacial interactions than avidin (27,28). More potent mutations are required to weaken this strong interface interaction.…”

“…Filtration-Observation of 100% monomerization of the streptavidin mutein using a non-boiled sample for SDS-PAGE does not always truly reflect its existence in the monomeric state in solution because SDS can promote subunit dissociation (27,28). The apparent molecular masses of the purified wild-type streptavidin and the three muteins (M2, M4, and AK) were estimated by gel filtration (Supplemental Fig.…”

Polydispersity as a Parameter for Indicating the Thermal Stability of Proteins by Dynamic Light Scattering

“…Other methods have been developed to evaluate the protein-ligand interaction in solution. Isothermal titration calorimetry (ITC) and differential scanning calorimetry (DSC) [10][11][12][13][14] have been used for evaluating protein function. Such calorimetric techniques measure the energy difference between a protein and its complex with a specific ligand.…”

Improvement of the thermal stability of streptavidin immobilized on magnetic beads by the construction of a mixed poly(ethylene glycol) tethered-chain layer

“…13 Therefore, we used heat treatment at 75 1C in this study. Before the incubation of the magnetic beads with the biotinylated ssDNA solutions, PEG/SA-MB and SA-MB were heated in an iCycler (Bio-Rad Laboratories, Inc., Hercules, CA, USA).…”

“…It is thought that heat-induced inactivation occurs because of the dissociation of intramolecular protein interactions. 13 Here, we report the preventative effect of a densely packed PEG layer against the heat-induced inactivation of streptavidin-immobilized magnetic beads (SA-MB). SA-MB co-immobilized with a PEG tethered-chain layer (PEG/SA-MB) were constructed by the co-immobilization of oligoamine-terminated PEGs onto SA-MB, and their biotin-binding efficiency was evaluated by measuring the amount of biotinylated single-stranded DNA (ssDNA) captured by the modified SA-MB.…”