Structure, conformational stability, and enzymatic properties of acylphosphatase from the hyperthermophile Sulfolobus solfataricus

The structure of AcP from the hyperthermophilic archaeon Sulfolobus solfataricus has been determined by 1H‐NMR spectroscopy and X‐ray crystallography. Solution and crystal structures (1.27 Å resolution, R‐factor 13.7%) were obtained on the full‐length protein and on an N‐truncated form lacking the f...

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Published in:Proteins, structure, function, and bioinformatics structure, function, and bioinformatics, 2006-01, Vol.62 (1), p.64-79
Main Authors: Corazza, Alessandra, Rosano, Camillo, Pagano, Katiuscia, Alverdi, Vera, Esposito, Gennaro, Capanni, Cristina, Bemporad, Francesco, Plakoutsi, Georgia, Stefani, Massimo, Chiti, Fabrizio, Zuccotti, Simone, Bolognesi, Martino, Viglino, Paolo
Format: Article
Language:English
Subjects:
1H NMR spectroscopy
Acid Anhydride Hydrolases - chemistry
Acid Anhydride Hydrolases - metabolism
Acylphosphatase
Archaeal Proteins - chemistry
Archaeal Proteins - metabolism
Enzyme Stability
Hydrogen-Ion Concentration
Kinetics
Protein Conformation
protein structure
protein thermostability
Scattering, Radiation
Sulfolobus - enzymology
Sulfolobus solfataricus
Sulfolobus solfataricus acylphosphatase
Thermodynamics
X-ray crystallography
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Summary:The structure of AcP from the hyperthermophilic archaeon Sulfolobus solfataricus has been determined by 1H‐NMR spectroscopy and X‐ray crystallography. Solution and crystal structures (1.27 Å resolution, R‐factor 13.7%) were obtained on the full‐length protein and on an N‐truncated form lacking the first 12 residues, respectively. The overall Sso AcP fold, starting at residue 13, displays the same βαββαβ topology previously described for other members of the AcP family from mesophilic sources. The unstructured N‐terminal tail may be crucial for the unusual aggregation mechanism of Sso AcP previously reported. Sso AcP catalytic activity is reduced at room temperature but rises at its working temperature to values comparable to those displayed by its mesophilic counterparts at 25–37°C. Such a reduced activity can result from protein rigidity and from the active site stiffening due the presence of a salt bridge between the C‐terminal carboxylate and the active site arginine. Sso AcP is characterized by a melting temperature, Tm, of 100.8°C and an unfolding free energy, ΔG U‐FH 2O, at 28°C and 81°C of 48.7 and 20.6 kJ mol−1, respectively. The kinetic and structural data indicate that mesophilic and hyperthermophilic AcP's display similar enzymatic activities and conformational stabilities at their working conditions. Structural analysis of the factor responsible for Sso AcP thermostability with respect to mesophilic AcP's revealed the importance of a ion pair network stabilizing particularly the β‐sheet and the loop connecting the fourth and fifth strands, together with increased density packing, loop shortening and a higher α‐helical propensity. Proteins 2006. © 2005 Wiley‐Liss, Inc.
ISSN:0887-3585
1097-0134
DOI:10.1002/prot.20703