Open AccessMechanistic Investigation of N-Homocysteinylation-Mediated Protein−Gold Nanoconjugate Assembly
Author(s) -
Arther T. Gates,
Leonard Moore,
Monica R. Sylvain,
Christina M. Jones,
Mark Lowry,
Bilal ElZahab,
James W. Robinson,
Robert M. Strongin,
Isiah M. Warner
Publication year - 2009
Publication title -
langmuir
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.042
H-Index - 333
eISSN - 1520-5827
pISSN - 0743-7463
DOI - 10.1021/la900798q
Subject(s) - chemistry , denaturation (fissile materials) , circular dichroism , colloidal gold , dynamic light scattering , covalent bond , biophysics , bovine serum albumin , surface plasmon resonance , human serum albumin , globular protein , nanoparticle , stereochemistry , biochemistry , nanotechnology , organic chemistry , nuclear chemistry , materials science , biology
Herein we report the use of protein-gold nanoconjugate (PGNs) as probes for elucidating mechanistic events involved in protein homocystamide detection with gold nanoparticles (GNPs), as was previously reported by our laboratory. Three different PGN probes are synthesized by direct adsorption of cytochrome c, albumin, or human serum onto citrate-capped GNPs. The PGNs are subsequently purified and treated to confer N-homocysteinylation. Individual PGN systems are evaluated to assess the effect of modification on (1) surface plasmon resonance (SPR), (2) protein structural conformation, and (3) assembly-association. The degree of PGN assembly and colorimetric signal observed postmodification varies based on the type of conjugated protein. For example, results of time-resolved dynamic light scattering studies indicate that modification of cytochrome c-PGNs yields rapid formation of macroscopic nanoparticle assemblies that eventually precipitate from solution. In contrast, albumin and human serum PGNs exhibit higher stability toward modification. Additionally, findings from circular dichroism studies indicate significant modification-induced denaturation, which is what may initiate assembly via electrosteric destabilization of PGNs. The results of electrophoretic studies appear to confirm that the process of N-homocysteinylation-mediated PGN assembly culminates in covalent interparticle association by disulfide cross-linking among modified proteins.