Tolerance of a protein to multiple polar-to-hydrophobic surface substitutions.


Abstract

Hydrophobic substitutions at solvent-exposed positions in two alpha-helical regions of the bacteriophage P22 Arc repressor were introduced by combinatorial mutagenesis. In helix A, hydrophobic residues were tolerated individually at each of the five positions examined, but multiple substitutions were poorly tolerated as shown by the finding that mutants with more than two additional hydrophobic residues were biologically inactive. Several inactive helix A variants were purified and found to have reduced thermal stability relative to wild-type Arc, with a rough correlation between the number of polar-to-hydrophobic substitutions and the magnitude of the stability defect. Quite different results were obtained in helix B, where variants with as many as five polar-to-hydrophobic substitutions were found to be biologically active and one variant with three hydrophobic substitutions had a t(m) 6 degrees C higher than wild-type. By contrast, a helix A mutant with three similar polar-to-hydrophobic substitutions was 23 degrees C less stable than wild-type. Also, one set of three polar-to-hydrophobic substitutions in helix B was tolerated when introduced into the wild-type background but not when introduced into an equally active mutant having a nearly identical structure. Context effects occur both when comparing different regions of the same protein and when comparing the same region in two different homologues. Study holds ProTherm entries: 5457, 5458 Extra Details: additive : EDTA(0.2 mM), aggregation; Arc repressor; combinatorial mutagenesis;,denatured state; protein folding; thermal stability

Submission Details

ID: n4f89U4o3

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:30 p.m.

Version: 1

Publication Details
Cordes MH;Sauer RT,Protein Sci. (1999) Tolerance of a protein to multiple polar-to-hydrophobic surface substitutions. PMID:10048325
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
1ARQ 1994-01-31 RELAXATION MATRIX REFINEMENT OF THE SOLUTION STRUCTURE OF THE ARC REPRESSOR
1ARR 1994-01-31 RELAXATION MATRIX REFINEMENT OF THE SOLUTION STRUCTURE OF THE ARC REPRESSOR
1QTG 1999-07-12 AVERAGED NMR MODEL OF SWITCH ARC, A DOUBLE MUTANT OF ARC REPRESSOR
1B28 1999-11-03 ARC REPRESSOR MYL MUTANT FROM SALMONELLA BACTERIOPHAGE P22
1NLA 2003-03-18 Solution Structure of Switch Arc, a Mutant with 3(10) Helices Replacing a Wild-Type Beta-Ribbon
1U9P 2005-02-15 1.9 Permuted single-chain Arc
1BAZ 1998-06-17 1.9 ARC REPRESSOR MUTANT PHE10VAL
1MYL 1995-01-26 2.4 SUBSTITUTING HYDROPHOBIC RESIDUES FOR A BURIED SALT BRIDGE ENHANCES PROTEIN STABILITY BUT DOES NOT REDUCE CONFORMATIONAL SPECIFICITY
1MYK 1995-01-26 2.4 CRYSTAL STRUCTURE, FOLDING, AND OPERATOR BINDING OF THE HYPERSTABLE ARC REPRESSOR MUTANT PL8
1BDT 1999-02-16 2.5 WILD TYPE GENE-REGULATING PROTEIN ARC/DNA COMPLEX
1PAR 1994-07-31 2.6 DNA RECOGNITION BY BETA-SHEETS IN THE ARC REPRESSOR-OPERATOR CRYSTAL STRUCTURE
1BDV 1999-01-06 2.8 ARC FV10 COCRYSTAL

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
100.0 Transcriptional repressor arc P03050 RARC_BPP22