Comparison of the three-dimensional protein and nucleotide structure of the FAD-binding domain of p-hydroxybenzoate hydroxylase with the FAD- as well as NADPH-binding domains of glutathione reductase.

The chain fold of the FAD-binding domain of p-hydroxybenzoate hydroxylase resembles the chain folds of the two nucleotide-binding domains of glutathione reductase. This fold consists of a four-stranded parallel beta-sheet sandwiched between a three-stranded antiparallel beta-sheet and alpha-helices. The nucleotides bind in similar positions relative to this chain fold. The best superposition of the folds has been established and geometrically quantified, giving rise to an equivalencing scheme for 110 residue positions, of which only four residues are identical in all three domains. It is discussed whether this chain fold is also present in a number of other FAD-binding proteins with known sequence. After the second strand of the parallel beta-sheet both FAD-binding domains contain long chain excursions, which make intimate contacts to rather distant parts of the respective molecules. In the environment of the isoalloxazine rings we observe interesting similarities. In both enzymes the si-face of this ring is covered by polypeptide, and only the re-face is accessible for the cofactor NADPH. Furthermore, there is a long alpha-helix in each enzyme, which points with its N-terminal start to the O-2 alpha region of isoalloxazine. These helices are spatially in the same position with respect to the isoalloxazine ring but are at quite different positions along the polypeptide chain. Since they can stabilize a negative charge around O-2 alpha, they may be important for the catalytic processes.