SCieNtifiC REPORTS | (2018) 8:3420 | DOI:10.1038/s41598-018-21571-y
Luminescent and paramagnetic
properties of nanoparticles shed
light on their interactions with
Giuditta Dal Cortivo
, Gabriel E. Wagner
, Paolo Cortelletti
, Krishna Mohan Padmanabha Das
, Adolfo Speghini
, Daniele Dell’Orco
& N. Helge Meyer
Nanoparticles have been recognized as promising tools for targeted drug-delivery and protein
therapeutics. However, the mechanisms of protein-nanoparticle interaction and the dynamics
underlying the binding process are poorly understood. Here, we present a general methodology for
the characterization of protein-nanoparticle interaction on a molecular level. To this end we combined
biophysical techniques including nuclear magnetic resonance (NMR), circular dichroism (CD), resonance
energy transfer (RET) and surface plasmon resonance (SPR). Particularly, we analyzed molecular
mechanisms and dynamics of the interaction of CaF
nanoparticles with the prototypical calcium
sensor calmodulin (CaM). We observed the transient formation of an intermediate encounter complex
involving the structural region linking the two domains. Specic interaction of CaM with CaF
driven by the N-terminal EF-hands, which seem to recognize Ca
on the surface of the nanoparticle. We
conclude that CaF
NP-CaM interaction is fully compatible with potential applications in nanomedicine.
Overall, the methods presented in this work can be extended to other systems and may be useful to
quantitatively characterize structural and dynamic features of protein-NP interactions with important
implications for nanomedicine and nano-biotechnology.
Nanoparticles (NPs) receive increasing attention in biomedical applications and are intensively discussed as
promising drug delivery systems in disease treatment
. Due to their high degree of specicity and minimal side
eects they are of particular interest as they can potentially be used with protein therapeutics in complex diseases.
However, the mechanisms of protein-nanoparticle interactions are not fully understood.
Recently, it has been shown that CaF
NPs can specically bind calcium sensor proteins including recoverin
guanylate cyclase-activating protein 1 (GCAP1)
and calmodulin (CaM)
. In these latter cases, binding to NPs
occurs at physiological concentrations in a fully reversible manner and most importantly, it does not alter sec-
ondary or tertiary structure of the protein
. e fact that the surface-bound protein remains structurally and
functionally intact suggests that CaF
NPs might be considered as specic carriers for calcium sensors including
CaM and GCAP1, and that exploiting the high surface-to-volume ratio typical of the nanoscale could constitute a
general strategy for protein replacement-therapy in the case of disease-associated mutant proteins
CaM is a prototypical calcium sensor protein, which is highly conserved and ubiquitous in eukaryotic cells.
It comprises four EF-hands (EF1-4), each containing a functional calcium binding motif and arranged in two
domains, termed C-terminal and N-terminal lobe. As Ca
ions act as important second messenger, CaM is
involved in many physiological processes including cell motility, proliferation, apoptosis, cytoskeleton remod-
eling, metabolic homeostasis, ion transport and protein folding
. Focusing on CaM-dependent biochemical
Department of Neurosciences, Biomedicine and Movement Sciences, Section of Biological Chemistry, Strada
le Grazie 8, Verona, Italy.
Institute of Hygiene, Microbiology and Environmental Medicine, Medical University of
Graz, Neue Stiftingtalstraße 6, 8010, Graz, Austria.
Nanomaterials Research Group, Department of Biotechnology,
University of Verona and INSTM, UdR Verona, Strada Le Grazie 15, Verona, Italy.
Institute of Molecular Biosciences,
University of Graz, Humboldtstraße 50/3, 8010, Graz, Austria.
Institute of Chemistry, University of Graz, Heinrichstr.
28, 8010, Graz, Austria.
Department of Human Medicine and Department of Neuroscience, University of Oldenburg,
Carl-von-Ossietzky-Str. 9-11, 26131, Oldenburg, Germany. Correspondence and requests for materials should be
addressed to D.D. (email: firstname.lastname@example.org) or N.H.M. (email: email@example.com)
Received: 14 November 2017
Accepted: 6 February 2018
Published: xx xx xxxx