The conformational stability and thermodynamics of Fur A (ferric uptake regulator) from Anabaena sp. PCC 7119

¹ Universidad de Zaragoza
² Universidad Miguel Hernandez
³ Molfsoft
⁴ University of Miguel Hernandez

^* To whom correspondence should be addressed. E-mail: jlneira{at}umh.es.

Submitted on May 4, 2005
Revised on June 6, 2005
Accepted on 22 August 2005

	Abstract

Fur (ferric uptake regulator) is a key bacterial protein that regulates iron acquisition and its storage, and modulates the expression of genes involved in the response to different environmental stresses. Although the protein is involved in several regulation mechanisms, and members of the Fur family have been identified in pathogen organisms, the stability and thermodynamic characterization of a Fur protein have not been described. In this work, the stability, thermodynamics and structure of the functional dimeric Fur A from Anabaena sp. PCC 7119 were studied by using computational methods and different biophysical techniques, namely, circular dichroism (CD), fluorescence, Fourier transform infrared (FTIR) and nuclear magnetic resonance (NMR) spectroscopies. The structure, as monitored by CD and FTIR, was composed of a 40 % of -helix. Chemical-denaturation experiments indicated that Fur A folded via a two-state mechanism, but its conformational stability was small with a value of G = 5.3 ± 0.3 kcal mol-1 at 298 K. Conversely, Fur A was thermally a highly stable protein. The high melting temperature (Tm = 352 ± 5 K), despite its moderate conformational stability, can be ascribed to its low heat capacity change upon unfolding, Cp, which had a value of 0.8 ± 0.1 kcal mol-1 K-1. This small value is probably due to burial of polar residues in the Fur A structure. This feature can be used for the design of mutants of Fur A with impaired DNA-binding properties.

Key Words: Fur protein, chemical-denaturations, heat capacity, protein folding, protein stability