Residue-specific immobilization of protein molecules by size-selected clusters.
The atomic force microscope (AFM), operating in contact mode, has been employed in buffer solution to study two proteins; (i) green fluorescent protein (GFP), from the hydromedusan jellyfish Aequorea victoria; and (ii) human oncostatin M (OSM), in the presence of size-selected gold nanoclusters pinned on to a highly oriented pyrolytic graphite substrate. The AFM images have revealed immobilization of single molecules of OSM, which are strongly bound to the gold nanoclusters. Conversely, no strong immobilization has been observed for the GFP, as these molecules were easily displaced by the scanning tip. The contrasting behaviour of the two proteins can be explained by the exposed molecular surface area of their cysteine residues as modelled on the basis of their respective X-ray crystallographic data structures. GFP contains two cysteine residues, but neither is readily available to chemisorb on the gold clusters, because the cysteines are largely inaccessible from the surface of the protein. In contrast, OSM has a total of five cysteine residues, with different degrees of accessibility, which make the protein amenable to anchoring on the nanoclusters. Statistical analysis of the height of the OSM molecules bound to the nanoclusters is in accordance with crystallographic data, and suggests various configurations of the proteins on the clusters, associated with the presence of different cysteine anchoring sites. These results suggest that the three-dimensional conformation of protein molecules is preserved when they are chemisorbed to size-selected gold clusters, thus opening a new route towards oriented immobilization of individual protein molecules.
Prisco
Umberto
U
Nanoscale Physics Research Laboratory, School of Physics and Astronomy, The University of Birmingham, Edgbaston, Birmingham B15 2TT, UK.
Leung
Carl
C
Xirouchaki
Chrisa
C
Jones
Celine H
CH
Heath
John K
JK
Palmer
Richard E
RE
D016207:Cytokines / Q000737:chemistry* / Q000648:ultrastructure; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000648:ultrastructure; D006801:Humans; D018625:Microscopy, Atomic Force; D008958:Models, Molecular; D008970:Molecular Weight; D053683:Oncostatin M; D010446:Peptide Fragments / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry* / Q000648:ultrastructure
D016207:Cytokines; C508351:OSM protein, human; D010446:Peptide Fragments; D011506:Proteins; D053683:Oncostatin M; D049452:Green Fluorescent Proteins