The influence of cross-linking on protein-protein interactions in a marine adhesive: the case of two byssus plaque proteins from the blue mussel.
The interaction between two proteins, Mefp-1 and Mefp-2, from the byssal plaque of the blue mussel, Mytilus edulis, was investigated using a quartz crystal microbalance with dissipation monitoring (QCM-D) technique. The challenge in using a surface-sensitive technique to investigate the interaction between two strongly adhesive proteins was met by coupling a biotinylated version of one of the proteins (b-Mefp-1) to an inert two-dimensional arrangement of streptavidin (SA) formed on top of a biotin-doped supported phospholipid bilayer. The interaction between Mefp-1 and Mefp-2 was further investigated by addition of Mefp-2 to SA-coupled b-Mefp-1, where the latter was either in the native state or cross-linked using sodium periodate (NaIO(4)), Cu(2+), or mushroom tyrosinase. With this coupling strategy it is shown that a requirement for attraction between the two proteins is that tyrosinase is used as the cross-linking agent of b-Mefp-1. By inhibiting the enzymatic activity of tyrosinase it is also shown that enzymatic activity is required for both efficient binding of tyrosinase to SA-coupled b-Mefp-1 as well as for the subsequent binding of Mefp-2. In contrast, spontaneous adsorption of Mefp-1 to a methyl-terminated (thiolated) gold surface followed by addition of Mefp-2 results in binding of Mefp-2 for all cross-linking agents. This suggests that cross-linking of Mefp-1 adsorbed on a solid surface induces structural changes in the adsorbed protein layer, resulting in exposure of free surface patches on which Mefp-2 binds.
Fant
Camilla
C
Department of Cell and Molecular Biology, Göteborg University, Box 462, SE-405 30 Göteborg, Sweden. camilla.fant@gmm.gu.se
Elwing
Hans
H
Höök
Fredrik
F
D000818:Animals; D049872:Bivalvia / Q000737:chemistry*; D003432:Cross-Linking Reagents / Q000138:chemical synthesis; D004295:Dihydroxyphenylalanine / Q000737:chemistry; D008081:Liposomes; D008722:Methods; D008970:Molecular Weight; D009991:Oscillometry; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry*
D003432:Cross-Linking Reagents; D008081:Liposomes; D011506:Proteins; C037403:adhesive protein, mussel; D004295:Dihydroxyphenylalanine