Filling small, empty protein cavities: structural and energetic consequences.


Abstract

Most proteins contain small cavities that can be filled by replacing cavity-lining residues by larger ones. Since shortening mutations in hydrophobic cores tend to destabilize proteins, it is expected that cavity-filling mutations may conversely increase protein stability. We have filled three small cavities in apoflavodoxin and determined by NMR and equilibrium unfolding analysis their impact in protein structure and stability. The smallest cavity (14 A3) has been filled, at two different positions, with a variety of residues and, in all cases, the mutant proteins are locally unfolded, their structure and energetics resembling those of an equilibrium intermediate of the thermal unfolding of the wild-type protein. In contrast, two slightly larger cavities of 20 A3 and 21 A3 have been filled with Val to Ile or Val to Leu mutations and the mutants preserve both the native fold and the equilibrium unfolding mechanism. From the known relationship, observed in shortening mutations, between stability changes and the differential hydrophobicity of the exchanged residues and the volume of the cavities, the filling of these apoflavodoxin cavities is expected to stabilize the protein by approximately 1.5 kcal mol(-1). However, both urea and thermal denaturation analysis reveal much more modest stabilizations, ranging from 0.0 kcal mol(-1) to 0.6 kcal mol(-1), which reflects that the accommodation of single extra methyl groups in small cavities requires some rearrangement, necessarily destabilizing, that lowers the expected theoretical stabilization. As the size of these cavities is representative of that of the typical small, empty cavities found in most proteins, it seems unlikely that filling this type of cavities will give rise to large stabilizations. Study holds ProTherm entries: 22077, 22078, 22079, 22080, 22081, 22082, 22083, 22084, 22085, 22086, 22087, 22088, 22089, 22090, 22091, 22092, 22093, 22094, 22095, 22096, 22097, 22098, 22099, 22100, 22101, 22102, 22103, 22104, 22105, 22106, 22107, 22108, 22109, 22110, 22111, 22112 Extra Details: apo form; Native to Intermediate; Global fitting of the four thermal unfolding curves recorded for each mutant protein (fluorescencel, far-UV CD, near-UV CD, and near-UV Abs). protein cavity; protein stability; protein intermediate; protein folding; van der Waals

Submission Details

ID: P9vDrB3j

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:53 p.m.

Version: 1

Publication Details
Bueno M;Cremades N;Neira JL;Sancho J,J. Mol. Biol. (2006) Filling small, empty protein cavities: structural and energetic consequences. PMID:16563433
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
2KQU 2010-06-02 F98N apoflavodoxin from Anabaena PCC 7119
5LJP 2017-08-02 1.1 E20K/I59A/E72K/I92A/D126K/A142V FLAVODOXIN FROM ANABAENA
1OBO 2003-04-24 1.2 W57L flavodoxin from Anabaena
1RCF 1995-01-26 1.4 STRUCTURE OF THE TRIGONAL FORM OF RECOMBINANT OXIDIZED FLAVODOXIN FROM ANABAENA 7120 AT 1.40 ANGSTROMS RESOLUTION
2V5V 2007-10-16 1.88 W57E Flavodoxin from Anabaena
3ESZ 2009-02-10 1.94 K2AK3A Flavodoxin from Anabaena
2V5U 2007-10-16 1.99 I92A FLAVODOXIN FROM ANABAENA
1FLV 1993-10-31 2.0 STRUCTURE OF THE OXIDIZED LONG CHAIN FLAVODOXIN FROM ANABAENA 7120 AT 2 ANGSTROMS RESOLUTION
1FTG 1996-12-23 2.0 STRUCTURE OF APOFLAVODOXIN: CLOSURE OF A TYROSINE/TRYPTOPHAN AROMATIC GATE LEADS TO A COMPACT FOLD
1QHE 1999-05-20 2.0 ENERGETICS OF A HYDROGEN BOND (CHARGED AND NEUTRAL) AND OF A CATION-PI INTERACTION IN APOFLAVODOXIN
1DX9 2000-04-10 2.05 W57A Apoflavodoxin from Anabaena
1OBV 2003-04-24 2.1 Y94F flavodoxin from Anabaena
3ESX 2009-02-10 2.31 E16KE61KD126KD150K Flavodoxin from Anabaena
3ESY 2009-02-10 2.39 E16KE61K Flavodoxin from Anabaena

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
100.0 Flavodoxin P0A3E0 FLAV_NOSSO
100.0 Flavodoxin P0A3D9 FLAV_NOSS1