ROLE OF AROMATIC RESIDUES IN STABILIZATION OF THE [FE4S4] CLUSTER IN HIGH-POTENTIAL IRON PROTEINS (HIPIPS) - PHYSICAL CHARACTERIZATION AND STABILITY STUDIES OF TYR-19 MUTANTS OF CHROMATIUM-VINOSUM HIPIP

The functional role of residue Tyr-19 of Chromatium vinosum HiPIP has been evaluated by site-directed mutagenesis experiments. The stability of the [Fe4S4] cluster prosthetic center is sensitive to side-chain replacements. Polar residues result in significant instability, while nonpolar residues (especially with aromatic side chains) maintain cluster stability. Two-dimensional NMR data of native and mutant HiPIPs are consistent with a model where Tyr-19 serves to preserve the structural rigidity of the polypeptide backbone, thereby maintaining a hydrophobic barrier for exclusion of water from the cluster cavity. Solvent accessibility results in more facile oxidation of the cluster by atmospheric oxygen, with subsequent rapid hydrolysis of the [Fe4S4](3+) core.