STRUCTURE DETERMINATION OF THE HUMAN PROTECTIVE PROTEIN - TWOFOLD AVERAGING REVEALS THE 3-DIMENSIONAL STRUCTURE OF A DOMAIN WHICH WAS ENTIRELY ABSENT IN THE INITIAL MODEL

Mutations in the human 'protective protein' result in the human lysoso mal storage disease galactosialidosis. The structure of the human 'pro tective protein' has been determined using X-ray crystallography to a resolution of 2.2 Angstrom. Initial phases were obtained from molecula r replacement calculations. A very partial search model comprising 30% of the scattering mass, was constructed from the atomic model of the wheat serine carboxypeptidase. This truncated probe was used to find t he position of two monomers in the asymmetric unit. Subsequently, 'boo tstrapping' cycles, consisting of twofold averaging and model expansio n, retrieved the electron density for residues initially missing. In p articular, it proved possible to add a domain (more than 110 residues) to the initial partial search model. In total, 314 residues per asymm etric unit were added to the 588 residues of the initial model. Factor s contributing to our success are discussed.