- vol.42, pp.331-336, 2000
C-1027 is a potent antitumor antibiotic, in which a nonprotein chromophore is tightly and specifically bound to an apoprotein. The chromophore, which has an endiyne structure and is responsible for DNA cleavage, is very labile when isolated, but greatly stabilized through binding to the apoprotein. Their binding structure and stabilizing interactions are very interesting problems in terms of molecular recognition and protein transport. The 3D structure of C-1027 apoprotein was determined by the X-PLOR calculation using 1539 experimental restrains derived from NMR spectroscopy. The apoprotein has three antiparallel β-sheets, and the hydrophobic pocket is formed by four-stranded β-sheet (DCHI) and two loops (residues 75-79, 97-100)(Fig.3). The overall shape of the apoprotein is quite similar to those of neocarzinostatin (NCS) and actinoxanthin (AXN). The binding structure of C-1027 complex was calculated based on 1539 NMR-derived constraints which include 38 intermolecular restraints between the aromatized chromophore and the apoprotein. The aromatized chromophore is bound to the hydrophobic pocket of the apoprotein. The benzodihydropentalene core locates in the center of the pocket with its molecular plane almost perpendicular to the bottom of the pocket. The β-tyrosine unit locates on the left side of the core, and both benzoxazine and aminosugar moieties on the right side (Fig.4). The hydrophobic interaction is most likely the major binding interaction between the apoprotein and the aromatized chromophore. Moreover, the 18-amino group of the chromophore locates in the proximity of either the carboxylate of Asp101 or the imidazole ring of His104, which indicates there could be a salt-bridge or a hydrogen bond type interaction between the aromatized chromophore and these side chains. To confirm the predicted binding interactions, we made several mutant apoproteins whose specific amino acid residue is replaced with the amino acid of different type and examined their binding abilities for the aromatized chromophore by NMR. The results obtained will be discussed.