Mechanistic Insights into Ultrasmall Gold Nanoparticle-Protein Interactions through Measurement of Binding Kinetics

Author(s)

Ferreira, Rodrigo S. ^[1] ; Lira, Andre L. ^[1] ; Torquato, Ricardo J. S. ^[1] ; Schuck, Peter ^[2] ; Sousa, Alioscka A. ^[1]

Affiliation

^[1] Univ Fed Sao Paulo, Dept Biochem, BR-04044020 Sao Paulo, SP - Brazil

^[2] Natl Inst Biomed Imaging & Bioengn, NIH, Bethesda, MD 20892 - USA

• Document type: Journal article
• Source: Journal of Physical Chemistry C; v. 123, n. 46, p. 28450-28459, NOV 21 2019.
• Web of Science Citations: 0

Abstract

Gold nanoparticles (NPs) in the ultrasmall size regime provide a new paradigm in the way that nanomaterials can be used to regulate protein structure and function. However, the rational design of ultrasmall NPs as viable synthetic effectors of protein function requires detailed quantitative understanding of their biomolecular interactions. Herein, we focused on the kinetics of NP-protein complexation-an often neglected factor in studies at the bio-nanointerface-to gain molecular-level insights into mechanisms of interactions. The protein alpha-thrombin and ultrasmall gold NPs coated with p-mercaptobenzoic acid (AuMBA) and glutathione (AuGSH) were used as model systems in our studies. Binding kinetics was quantified by surface plasmon resonance (SPR) biosensing and stopped-flow spectroscopy. The results revealed strong and weak interactions of AuMBA and AuGSH toward thrombin (K-D similar to 30 nM and 20 mu M, respectively), as well as ``fast{''} and ``slow{''} association kinetics (k(on) similar to 10(6)-10(7) and 10(4) M-1 s(-1) respectively). The significantly smaller k(on) for AuGSH implied the presence of a larger energy barrier along the association pathway, presumably related to the penalty required to remove interfacial ions. Analysis of dissociation reactions revealed that thrombin interactions with both NPs formed transient, weakly adhesive complexes characterized by short residence times (t(r) = 1/k(off) similar to 0.1-16 s). Interestingly, the reverse reactions were best described by multiple dissociation processes suggesting a heterogeneous population of complexes stabilized to different extent. (AU)

• FAPESP's process: 19/04372-6 - Interactions of ultrasmall gold nanoparticles with proteins and cells
• Grantee: Alioscka Augusto Sousa
• Support Opportunities: Regular Research Grants
• FAPESP's process: 16/25535-2 - Biointeractions of gold nanoparticles: influence of macromolecular crowding and soft interactions
• Grantee: Rodrigo da Silva Ferreira
• Support Opportunities: Scholarships in Brazil - Post-Doctoral