Probing the effects of cysteine residues on protein adsorption onto gold nanoparticles using wild-type and mutated GB3 proteins.
The role of cysteine residues in the protein binding kinetics and stability on gold nanoparticles (AuNP) was studied using AuNP localized surface plasmon resonance (LSPR) in combination with an organothiol (OT) displacement method. GB3, the third IgG-binding domain of protein G, was used to model protein-AuNP adsorption. While wild-type GB3 (GB30) contains no cysteine residues, bioengineered GB3 variants containing one (GB31) and two (GB32) cysteine residues were also tested. The cysteine content has no significant effect on GB3 binding kinetics with AuNPs, and most protein adsorption occurs within the first few seconds upon protein/AuNP mixing. However, the stability of GB3 on the AuNP surface against OT displacement depends strongly on the cysteine content and the age of the AuNP/GB3 mixture. The GB30 covered AuNPs can be completely destabilized and aggregated by OTs, regardless of the age of the GB30/AuNP mixtures. Long-time incubation of GB31 or GB32 with AuNPs can stabilize AuNPs against the OT adsorption inducted aggregation. This study indicates that multiple forces involved in the GB3/AuNP interaction, and covalent binding between cysteine and AuNP is essential for a stable protein/AuNP complex.
Department of Chemistry, Mississippi State University, Mississippi State, Mississippi 39762, USA.
Kumudu
K
Siriwardana
Ailin
A
Wang
Karthikeshwar
K
Vangala
Nicholas
N
Fitzkee
Dongmao
D
Zhang
D000327:Adsorption; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D003545:Cysteine / Q000737:chemistry; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006046:Gold / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D053768:Metal Nanoparticles / Q000737:chemistry*; D009154:Mutation; D011485:Protein Binding; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D013439:Sulfhydryl Reagents / Q000737:chemistry*; D020349:Surface Plasmon Resonance; D013499:Surface Properties
D001426:Bacterial Proteins; C034284:IgG Fc-binding protein, Streptococcus; D011994:Recombinant Proteins; D013439:Sulfhydryl Reagents; D006046:Gold; D003545:Cysteine