Role of the N-terminal region for the conformational stability of esterase 2 from Alicyclobacillus acidocaldarius

Abstract

of the N-terminal region for the conformational stability of esterase 2 from Alicyclobacillus acidocaldarius. Biophysical Chemistry, Elsevier, 2007, 127 (1-2) This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. A C C E P T E D M A N U S C R I P T ACCEPTED MANUSCRIPT A C C E P T E D M A N U S C R I P T ACCEPTED MANUSCRIPT

Molecular dynamics study of the conformational stability of esterase 2 from Alicyclobacillus acidocaldarius

“…For instance, it is not correct to identify one of the two domains simply with the so-called "cap" domain, recognized in the X-ray structure of EST2 and containing the protein N-terminal region [11]. Deletion of the first 35 residues of the polypeptide chain did not lead to a more cooperative thermal unfolding process, as could be expected, but to the loss of important interactions between the two domains that, as a consequence, behave in a much more independent manner [12].…”

“…Deconvolution of DSC traces showed the presence of two cooperative and coupled domains in both proteins, and allowed the determination of the thermodynamic parameter values characterizing the two component transitions [12]. One of the two domains is more stable with a denaturation temperature of 86 • C, common to both proteins, whereas the other domain has a denaturation temperature that is 81 • C for EST2, and 62 • C for EST2-36del [12]. Identification of the EST2 domains responsible of such behaviour was not straightforward in view of the structural features of the protein.…”

“…It was suggested that the external shell of EST2 structure corresponds to the less stable domain, while the core region of EST2 structure corresponds to the more stable domain [12]. In order to arrive at a more definitive identification of the two domains existing in the EST2 structure, molecular dynamics (MD) simulations at three different temperatures, namely 300, 400 and 500 K, in explicit water, have been performed on both EST2 and EST2-36del.…”

“…Circular dichroism, CD, and differential scanning calorimetry, DSC, measurements were performed to investigate in detail the thermal unfolding of EST2 and EST2-36del, showing that the temperature-induced unfolding of the two proteins is a reversible process that cannot be represented as a two-state N D transition [12]. Deconvolution of DSC traces showed the presence of two cooperative and coupled domains in both proteins, and allowed the determination of the thermodynamic parameter values characterizing the two component transitions [12].…”

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a b s t r a c t Circular dichroism and differential scanning calorimetry measurements showed that esterase 2 from the thermophilic microorganism Alicyclobacillus acidocaldarius, EST2, and its variant in which the first 35 residues have been deleted, EST2-36del, unfold reversibly on increasing temperature, and possess two cooperative and coupled domains [12]. Structural features of the ␣/␤ hydrolase fold of EST2, with nine ␣-helices packed against the central twisted ␤-sheet, do not allow a straightforward identification of these two cooperative and coupled domains.

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