Dimerization of a His117Gly azurin mutant by external addition of 1,omega-di(imidazol-1-yl)alkanes.
The possibility to construct non-covalently linked protein dimers was investigated by employing the His117Gly mutant of the Cu containing azurin and the bifunctional 1,omega-di(imidazol-1-yl)alkanes as linkers. The His117Gly mutation creates a gap in the coordination sphere of the metal through which the latter becomes accessible for externally added ligands. The bifunctional ligands gave rise to the formation of dimers provided the linker was sufficiently long, as in the case of 1,omega-di(imidazol-1-yl)pentane and -hexane; the butane linker only produced monomers. The binding of the azurin molecules to the bifunctional C5 and C6 linkers showed cooperativity, which is the result of the hydrophobic interaction of the aligned hydrophobic patches. The energy and surface area involved in this process have been estimated from the experimental data to be delta G is -1.3 to -2.1 kcal/mol and 65-105 A2. The implications for the study of electron transfer processes inside a protein matrix are indicated.
van Pouderoyen
G
G
Leiden Institute of Chemistry, Gorlaeus Laboratories, Leiden University, The Netherlands.
den Blaauwen
T
T
Reedijk
J
J
Canters
G W
GW
D000473:Alkanes / Q000378:metabolism*; D001400:Azurin / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D003300:Copper / Q000378:metabolism; D003432:Cross-Linking Reagents / Q000378:metabolism*; D019281:Dimerization; D004578:Electron Spin Resonance Spectroscopy; D007093:Imidazoles / Q000378:metabolism*; D008024:Ligands; D008958:Models, Molecular; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D011550:Pseudomonas aeruginosa / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013053:Spectrophotometry; D013816:Thermodynamics
D000473:Alkanes; D003432:Cross-Linking Reagents; D007093:Imidazoles; D008024:Ligands; D011994:Recombinant Proteins; D001400:Azurin; D003300:Copper