HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Moczygemba, C. Articles by Wittung-Stafshede, P. Search for Related Content PubMed PubMed Citation Articles by Moczygemba, C. Articles by Wittung-Stafshede, P. Social Bookmarking                   What's this?

High stability of a ferredoxin from the hyperthermophilic archaeon A. ambivalens: Involvement of electrostatic interactions and cofactors

¹ Chemistry Department, Tulane University, New Orleans, Louisiana 70118, USA
² Molecular and Cellular Biology Graduate Program, Tulane University, New Orleans, Louisiana 70118, USA
³ Instituto de Technologia Quimica e Biologica, Universidade Nova de Lisboa, 2780-156 Oeiras, Portugal

Reprint requests to: Pernilla Wittung-Stafshede, Chemistry Department, Tulane University, 6823 St. Charles Avenue, New Orleans, LA 70118-5698, USA; e-mail: pernilla{at}tulane.edu; fax: (504)-865-5596.

The ferredoxin from the thermophilic archaeon Acidianus ambivalens is a small monomeric seven-iron protein with a thermal midpoint (T_m) of 122°C (pH 7). To gain insight into the basis of its thermostability, we have characterized unfolding reactions induced chemically and thermally at various pHs. Thermal unfolding of this ferredoxin, in the presence of various guanidine hydrochloride (GuHCl) concentrations, yields a linear correlation between unfolding enthalpies (H[T_m]) and T_m from which an upper limit for the heat capacity of unfolding (C_P) was determined to be 3.15 ± 0.1 kJ/(mole • K). Only by the use of the stronger denaturant guanidine thiocyanate (GuSCN) is unfolding of A. ambivalens ferredoxin at pH 7 (20°C) observed ([GuSCN]_1/2 = 3.1 M; G_U[H₂O] = 79 ± 8 kJ/mole). The protein is, however, less stable at low pH: At pH 2.5, T_m is 64 ± 1°C, and GuHCl-induced unfolding shows a midpoint at 2.3 M (G_U[H₂O] = 20 ± 1 kJ/mole). These results support that electrostatic interactions contribute significantly to the stability. Analysis of the three-dimensional molecular model of the protein shows that there are several possible ion pairs on the surface. In addition, ferredoxin incorporates two iron–sulfur clusters and a zinc ion that all coordinate deprotonated side chains. The zinc remains bound in the unfolded state whereas the iron–sulfur clusters transiently form linear three-iron species (in pH range 2.5 to 10), which are associated with the unfolded polypeptide, before their complete degradation.

Keywords: Hyperthermophiles; thermostability; iron-sulfur proteins; protein unfolding

Abbreviations: G_U, unfolding free energy • T_m, melting temperature • Fd, ferredoxin • GuHCl, guanidine hydrochloride • H[T_m], enthalpy of unfolding • C_P, heat capacity of unfolding • GuSCN, guanidine thiocyanate • CD, circular dichroism

CiteULike    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				A. Sujak, N. J. M. Sanghamitra, O. Maneg, B. Ludwig, and S. Mazumdar Thermostability of Proteins: Role of Metal Binding and pH on the Stability of the Dinuclear CuA Site of Thermus thermophilus Biophys. J., October 15, 2007; 93(8): 2845 - 2851. [Abstract] [Full Text] [PDF]

				H.-X. Zhou Toward the Physical Basis of Thermophilic Proteins: Linking of Enriched Polar Interactions and Reduced Heat Capacity of Unfolding Biophys. J., December 1, 2002; 83(6): 3126 - 3133. [Abstract] [Full Text] [PDF]