A molecular mechanics and database analysis of the structural preorganization and activation of the chromophore-containing hexapeptide fragment in green fluorescent protein.
We propose that heterologous posttranslational chromophore formation in green fluorescent protein (GFP) occurs because the chromophore-forming amino acid residues 65SYG67 are preorganized and activated for imidazolinone ring formation. Based on extensive molecular mechanical conformational searching of the precursor hexapeptide fragment (64FSYGVQ69), we suggest that the presence of low energy conformations characterized by short contacts (approximately 3 A) between the carbonyl carbon of Ser65 and the amide nitrogen of Gly67 accounts for the initial step in posttranslational chromophore formation. Database searches showed that the tight turn required to establish the key short contact is a unique structural motif that is rarely found, except in other FSYG tetrapeptide sequences. Additionally, ab initio calculations demonstrated that an arginine side chain can hydrogen bond to the carbonyl oxygen of Ser65, activating this group for nucleophilic attack by the nearby lone pair of the Gly67 amide nitrogen. We propose that GFP chromophore-formation is initiated by a unique combination of conformational and electronic enhancements, identified by computational methods.
Branchini
B R
BR
Chemistry Department, Connecticut College, New London 06320, USA.
Lusins
J O
JO
Zimmer
M
M
D055598:Chemical Phenomena; D002627:Chemistry, Physical; D016208:Databases, Factual; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000737:chemistry*; D008958:Models, Molecular; D010446:Peptide Fragments / Q000737:chemistry; D011487:Protein Conformation
D008164:Luminescent Proteins; D010446:Peptide Fragments; D049452:Green Fluorescent Proteins