Role of a surface tryptophan in defining the structure, stability, and DNA binding of the hyperthermophile protein Sac7d.


Abstract

Sac7d is a small, chromatin protein from Sulfolobus acidocaldarius which induces a sharp kink in DNA with intercalation of valine and methionine side chains. The crystal structure of the protein-DNA complex indicates that a surface tryptophan (W24) plays a key role in DNA binding by hydrogen bonding to the DNA at the kink site. We show here that substitution of the solvent-exposed tryptophan with alanine (W24A) led to a significant loss in not only DNA binding affinity but also protein stability. The W24A substitution proved to be one of the most destabilizing surface substitutions in Sac7d. A global linkage analysis of the pH and salt dependence of stability indicated that the protein stability surface (DeltaG vs temperature, pH, and salt concentration) was lowered overall by 2 kcal/mol (from 0 to 100 degrees C, pH 0 to 7, and 0 to 0.3 M KCl). The lower free energy of unfolding could not be attributed to significant structural perturbations of surface electrostatic interactions. Residual dipolar coupling of partially aligned protein and the NMR solution structure of W24A confirmed that the surface substitution resulted in no significant change in structure. Stabilization of this hyperthermophile protein and its DNA complex by a surface cluster of hydrophobic residues involving W24 and the two intercalating side chains is discussed. Study holds ProTherm entries: 18641, 18642 Extra Details: DNA binding; hydrogen bonding; solvent-exposed; electrostatic interactions; hydrophobic residues DNA binding; hydrogen bonding; solvent-exposed; electrostatic interaction; hydrophobic residues

Submission Details

ID: xfcWUXoV

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:50 p.m.

Version: 1

Publication Details
Bedell JL;Edmondson SP;Shriver JW,Biochemistry (2005) Role of a surface tryptophan in defining the structure, stability, and DNA binding of the hyperthermophile protein Sac7d. PMID:15654747
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