1 research outputs found
Oriented Attachment of Recombinant Proteins to Agarose-Coated Magnetic Nanoparticles by Means of a β‑Trefoil Lectin Domain
Design of generic methods aimed at
the oriented attachment of proteins
at the interfacial environment of magnetic nanoparticles currently
represents an active field of research. With this in mind, we have
prepared and characterized agarose-coated maghemite nanoparticles
to set up a platform for the attachment of recombinant proteins fused
to the β-trefoil lectin domain LSL<sub>150</sub>, a small protein
that combines fusion tag properties with agarose-binding capacity.
Analysis of the agarose-coated nanoparticles by dynamic light scattering,
Fourier transform infrared spectroscopy, and thermogravimetric studies
shows that decoupling particle formation from agarose coating provides
better results in terms of coating efficiency and particle size distribution.
LSL<sub>150</sub> interacts with these agarose-coated nanoparticles
exclusively through the recognition of the sugars of the polymer,
forming highly stable complexes, which in turn can be dissociated
ad hoc with the competing sugar lactose. Characterization of the complexes
formed with the fusion proteins LSL-EGFP (LSL-tagged enhanced green
fluorescent protein from Aquorea victoria) and LSL-BTL2 (LSL-tagged lipase from Geobacillus
thermocatenolatus) provided evidence supporting a
topologically oriented binding of these molecules to the interface
of the agarose-coated nanoparticles. This is consistent with the marked
polarity of the β-trefoil structure where the sugar-binding
sites and the N- and C-terminus ends are at opposed sides. In summary,
LSL<sub>150</sub> displays topological and functional features expected
from a generic molecular adaptor for the oriented attachment of proteins
at the interface of agarose-coated nanoparticles