Concentration-dependent β-cyclodextrin-promoted refolding of gold nanoparticles-conjugated bovine serum albumin complexed with gemini surfactants with different spacer groups

This work demonstrates refolding of unfolded bioconjugated bovine serum albumin [(BSA, 10 mu M):gold nano particles (AuNPs, 0.0124 mu M)] using beta-cyclodextrin (beta-CD). Protein bioconjugates are first unfolded with 0.2 mM of each of gemini surfactants, 12-4-12,2Br and 12-8-12,2Br-that have hydrophobic spacer groups of different lengths. At this concentration of a gemini surfactant, protein bioconjugates remain in compressed form due to strong hydrophobic interactions imparted by highly hydrophobic microdomain within the core of the gemini micelles formed along the chain of a bioconjugate. The compressed protein bioconjugates get relaxed in presence of a low concentration of beta-CD which strips off surfactant molecules from bioconjugate-micelles complexes. Relaxed denatured protein bioconjugates then get refolded once remaining surfactant molecules are stripped off at a higher concentration of beta-CD. A step-wise refolding process of bioconjugated protein chains exactly opposite to the unfolding is noted. Binding isotherms are constructed using Forster's resonance energy transfer (FRET) and nanometal surface energy transfer (NSET) parameters calculated based on energy transfer process occurs between Trp residues and AuNPs. a-helix of protein bioconjugates remains in inverse relationship to beta-turns during refolding by beta-CD. Refolding is found to be more in presence of 12-8-12,2Br- than 12-4-12,2Br-. Secondary/ tertiary structural changes of refolded BSA, on being bioconjugated, have been determined through various spectroscopic analysis. An approach towards construing the refolding of denatured AuNPs-bioconjugates as potential materials and impacts of AuNPs on conformations of refolded protein is presented here.

Automated summary

This work demonstrates the reverse refolding process of unfolded bioconjugated proteins using beta-cyclodextrin (beta-CD). The compressed protein bioconjugates relax in the presence of a low concentration of beta-CD and then refold once remaining surfactant molecules are removed at a higher concentration of beta-CD. The secondary/tertiary structural changes of the refolded protein have been determined through spectroscopic analysis. This study provides insights into the protein folding process and its potential applications in materials science.