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
Additional Information

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P
 

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)


Relevant PDB Entries

Structure ID Release Date Resolution Structure Title
1XX8 2005-02-08 NMR Structure of the W24A Mutant of the Hyperthermophile Sac7d Protein
1SAP 1995-09-15 HYPERTHERMOPHILE PROTEIN, RELAXATION MATRIX REFINEMENT STRUCTURE
1XYI 2005-02-08 1.45 Hyperthermophile chromosomal protein Sac7d double mutant Val26Ala/Met29Ala in complex with DNA GCGATCGC
4CJ2 2014-05-21 1.5 Crystal structure of HEWL in complex with affitin H4
2XIW 2011-07-13 1.5 Crystal structure of the Sac7d-derived IgG1-binder C3-C24S
1WTO 2005-02-22 1.5 Hyperthermophile chromosomal protein SAC7D double mutant V26F/M29F in complex with DNA GCGATCGC
1BF4 1999-11-10 1.6 CHROMOSOMAL DNA-BINDING PROTEIN SSO7D/D(GCGAACGC) COMPLEX
1WTV 2005-02-22 1.6 Hyperthermophile chromosomal protein SAC7D single mutant M29A in complex with DNA GTAATTAC
1AZP 1999-01-13 1.6 HYPERTHERMOPHILE CHROMOSOMAL PROTEIN SAC7D BOUND WITH KINKED DNA DUPLEX
4CJ1 2014-05-21 1.63 Crystal structure of CelD in complex with affitin H3
1WTQ 2005-02-22 1.7 Hyperthermophile chromosomal protein SAC7D single mutant M29F in complex with DNA GTAATTAC
1WTR 2005-02-22 1.8 Hyperthermophile chromosomal protein SAC7D single mutant M29A in complex with DNA GCGATCGC
1WTP 2005-02-22 1.9 Hyperthermophile chromosomal protein SAC7D single mutant M29F in complex with DNA GCGA(UBr)CGC
1WD0 2004-08-03 1.9 Crystal structures of the hyperthermophilic chromosomal protein Sac7d in complex with DNA decamers
1AZQ 1999-01-13 1.94 HYPERTHERMOPHILE CHROMOSOMAL PROTEIN SAC7D BOUND WITH KINKED DNA DUPLEX
1WTX 2005-02-22 2.2 Hyperthermophile chromosomal protein SAC7D single mutant V26A in complex with DNA GTAATTAC
1CA6 2000-02-23 2.2 INTERCALATION SITE OF HYPERTHERMOPHILE CHROMOSOMAL PROTEIN SSO7D/SAC7D BOUND TO DNA
1WD1 2004-08-03 2.2 Crystal structures of the hyperthermophilic chromosomal protein Sac7d in complex with DNA decamers
1CA5 2000-02-23 2.2 INTERCALATION SITE OF HYPERTHERMOPHILE CHROMOSOMAL PROTEIN SSO7D/SAC7D BOUND TO DNA
1WTW 2005-02-22 2.2 Hyperthermophile chromosomal protein SAC7D single mutant V26A in complex with DNA GCGATCGC
1WVL 2005-08-02 2.6 Crystal Structure of Multimeric DNA-binding Protein Sac7d-GCN4 with DNA decamer

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
92.9 DNA-binding protein 7d F4B8X5 DN7C_ACIHW
91.4 DNA-binding protein 7d F4B9I5 DN7B_ACIHW
92.3 DNA-binding protein 7d P13125 DN7E_SULAC
100.0 DNA-binding protein 7d P13123 DN7D_SULAC