Abstract
BackgroundGold nanoparticles (AuNP) are extensively used as biophysical tools in the area of medicine and technology due to their distinct properties. However, vivid understanding of the consequences of biomolecule-nanomaterial interactions is still lacking. In this context, we explore the affect of conjugation of Gαi1 subunit (of heterotrimeric G-proteins) to AuNP and examine its consequences. We consider two bio-conjugation strategies covalent and non-covalent binding.ResultsAffinity of the AuNP to the Gαi1 is 7.58 × 10 12 M-1. AuNP conjugated Gαi1 exhibits altered kinetics of activation, non-covalent bio-conjugates displays retarded kinetics, up to 0.88 fold when GTPγS was used as ligand, of protein activation contrary to covalent conjugates which accelerates it to ~ 5 fold. Conjugation influence intrinsic Gαi1 GTPase function in conflicting modes. Non-covalent conjugation inhibits GTPase function (decrease in activity upto 0.8 fold) whilst covalent conjugation drastically accelerates it (12 fold increase in activity). Altered basal nucleotide uptake in both types of conjugates and GTPase function in non-covalent conjugate are almost comparable except for GTPase property of covalent conjugate. The effect is despite the fact that conjugation does not change global conformation of the protein.ConclusionThese findings provide clear evidence that nanoparticles, in addition to ‘passive interaction’ with protein (biomolecule), can interact “actively” with biomolecule and modify its function. This concept should be considered while engineering nanoparticle based delivery systems in medicine.
Highlights
Gold nanoparticles (AuNP) are extensively used as biophysical tools in the area of medicine and technology due to their distinct properties
Dihydrolipoic acid (DHLA) capped a dark-red solution of Gold nanoparticles (AuNPs) of hydrodynamic diameter ~ 6 nm was used in the entire study
(i) N-terminal covalent conjugation using Ethyl-3-[3-dimethylamino propyl] carbodiimide (EDC) chemistry: Site specific conjugation was achieved by forming a peptide bond between N-terminal primary amine of the protein and carboxylic acid groups of negatively charged AuNP utilizing 1-Ethyl-3-[3dimethylaminopropyl] carbodiimide, EDC, chemistry
Summary
Gold nanoparticles (AuNP) are extensively used as biophysical tools in the area of medicine and technology due to their distinct properties. Vivid understanding of the consequences of biomoleculenanomaterial interactions is still lacking In this context, we explore the affect of conjugation of Gαi subunit (of heterotrimeric G-proteins) to AuNP and examine its consequences. AuNP conjugated Gαi exhibits altered kinetics of activation, non-covalent bio-conjugates displays retarded kinetics, up to 0.88 fold when GTPγS was used as ligand, of protein activation contrary to covalent conjugates which accelerates it to ~ 5 fold. G proteins are ubiquitously expressed and despite the variety in their function and biochemical effects, their structures are very highly conserved. These properties of G proteins, make them very vital model systems for studying the effects of nanoparticles; an area that is fast gaining importance in biology and medicine
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