title: Crystallographic and spectroscopic characterizations of Sulfolobus solfataricus TrxA1 provide insights into the determinants of thioredoxin fold stability.
abstract: Structural characterizations of thioredoxins (Trxs) are important for their involvement in severe pathologies and for their stable scaffold. Here we report a combined structural and spectroscopic characterization of a Trx isolated from the hyperthermophilic archaeon Sulfolobus solfataricus (SsTrxA1). Thermal denaturation unveils that SsTrxA1 is endowed with a remarkable stability in the explored temperature range 50-105°C. The structure of the oxidized form of SsTrxA1 determined at 1.9Å resolution presents a number of peculiar features. Although the protein was crystallized in a slightly acid medium (pH 6.5) as many as ten intramolecular/intermolecular carboxyl-carboxylate interactions involving glutamic and aspartic acid side chains are found in three independent SsTrxA1 molecules present in the asymmetric unit. Surprisingly for a hyperthermostable protein, the structure of SsTrxA1 is characterized by the presence (a) of a very limited number of intramolecular salt bridges and (b) of a cavity nearby Cys52, a residue that is frequently a phenylananine in other members of the family. Chemical denaturation investigations carried out on SsTrxA1 and SsTrxA2 show that both proteins present a significant stability against guanidine hydrochloride, thus indicating that ionic interactions play a minor role in their stabilization. Compared to Trxs from mesophilic sources, SsTrxA1 displays a longer α-helix 1 and a shorter loop connecting this α-helix with β-strand 2. As these features are shared with Trxs isolated from thermophilic sources, the shortening of this loop may be a general strategy adopted to stabilize this fold. This feature may be exploited for the design of hyperthermostable Trx scaffolds.
authors.affiliation: Istituto di Biostrutture e Bioimmagini, CNR, Via Mezzocannone 16, I-80134 Napoli, Italy.
authors.forename: Luciana
authors.initials: L
authors.lastname: Esposito
authors.forename: Alessia
authors.initials: A
authors.lastname: Ruggiero
authors.forename: Mariorosario
authors.initials: M
authors.lastname: Masullo
authors.forename: Maria Rosaria
authors.initials: MR
authors.lastname: Ruocco
authors.forename: Anna
authors.initials: A
authors.lastname: Lamberti
authors.forename: Paolo
authors.initials: P
authors.lastname: Arcari
authors.forename: Adriana
authors.initials: A
authors.lastname: Zagari
authors.forename: Luigi
authors.initials: L
authors.lastname: Vitagliano
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000737:chemistry*; D002942:Circular Dichroism; D018360:Crystallography, X-Ray; D006860:Hydrogen Bonding; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011489:Protein Denaturation; D055550:Protein Stability; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D040681:Structural Homology, Protein; D048229:Sulfolobus solfataricus; D013879:Thioredoxins / Q000737:chemistry*
chemical_list: D019843:Archaeal Proteins; D013879:Thioredoxins