title: Role of pyridoxal 5'-phosphate in the structural stabilization of O-acetylserine sulfhydrylase.
abstract: Proteins belonging to the superfamily of pyridoxal 5'-phosphate-dependent enzymes are currently classified into three functional groups and five distinct structural fold types. The variation within this enzyme group creates an ideal system to investigate the relationships among amino acid sequences, folding pathways, and enzymatic functions. The number of known three-dimensional structures of pyridoxal 5'-phosphate-dependent enzymes is rapidly increasing, but only for relatively few have the folding mechanisms been characterized in detail. The dimeric O-acetylserine sulfhydrylase from Salmonella typhimurium belongs to the beta-family and fold type II group. Here we report the guanidine hydrochloride-induced unfolding of the apo- and holoprotein, investigated using a variety of spectroscopic techniques. Data from absorption, fluorescence, circular dichroism, (31)P nuclear magnetic resonance, time-resolved fluorescence anisotropy, and photon correlation spectroscopy indicate that the O-acetylserine sulfhydrylase undergoes extensive disruption of native secondary and tertiary structure before monomerization. Also, we have observed that the holo-O-acetylserine sulfhydrylase exhibits a greater conformational stability than the apoenzyme form. The data are discussed in light of the fact that the role of the coenzyme in structural stabilization varies among the pyridoxal 5'-phosphate-dependent enzymes and does not seem to be linked to the particular enzyme fold type.
authors.affiliation: Institute of Physical Sciences, Institute of Biochemical Sciences, and National Institute for the Physics of Matter, University of Parma, Parma 43100, Italy.
authors.forename: S
authors.initials: S
authors.lastname: Bettati
authors.forename: S
authors.initials: S
authors.lastname: Benci
authors.forename: B
authors.initials: B
authors.lastname: Campanini
authors.forename: S
authors.initials: S
authors.lastname: Raboni
authors.forename: G
authors.initials: G
authors.lastname: Chirico
authors.forename: S
authors.initials: S
authors.lastname: Beretta
authors.forename: K D
authors.initials: KD
authors.lastname: Schnackerz
authors.forename: T L
authors.initials: TL
authors.lastname: Hazlett
authors.forename: E
authors.initials: E
authors.lastname: Gratton
authors.forename: A
authors.initials: A
authors.lastname: Mozzarelli
mesh_terms: D003547:Cysteine Synthase / Q000737:chemistry*; D019791:Guanidine / Q000737:chemistry; D008027:Light; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular; D011487:Protein Conformation; D011489:Protein Denaturation; D011732:Pyridoxal Phosphate / Q000737:chemistry*; D012542:Scattering, Radiation; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet
chemical_list: D011732:Pyridoxal Phosphate; D003547:Cysteine Synthase; D019791:Guanidine