Conformational stability of Helicobacter pylori flavodoxin: fit to function at pH 5.


Abstract

Flavodoxin is an essential protein for Helicobacter pylori, a pathogen living in the very acidic environment of the gastric tract and responsible for several diseases. We report the conformational stability of the protein in neutral and acidic pH. The apoprotein remains native between pH 12 and 5 and adopts a monomeric molten globule conformation at more acidic pH values. The equilibrium unfolding in urea appears two-state for either conformation, but the native one coexists with a hidden equilibrium intermediate of very similar properties. The stability of H. pylori apoflavodoxin is higher than that of the Anabaena homologue throughout the entire pH interval, which may be related to better charge compensation. H. pylori apoflavodoxin is strongly stabilized by its FMN cofactor. A global analysis of apo- and holoflavodoxin equilibrium unfolding, with and without excess FMN, indicates that the cofactor only binds to the native state. Some physical-chemical properties of the protein may represent an adaptation to the acidic environment. Unlike the apoflavodoxin from Anabaena, which becomes highly insoluble at pH 5.0, that from H. pylori remains soluble to at least 40 microm. This fact, together with the high stability of the apoprotein at this low pH that can arise in the bacteria cytoplasm, seems useful to allow newly synthesized apoflavodoxin molecules to fold and remain soluble to accomplish cofactor binding, which in turn increases the stability. Also, whenever the cytoplasmic pH drops to 5, preexisting flavodoxin molecules will remain folded and soluble and will retain the FMN cofactor, thus remaining functional. Study holds ProTherm entries: 24782, 24783, 24784, 24785, 24786, 24787, 24788, 24789 Extra Details: conformational stability; intermediate; charge compensation; cofactor

Submission Details

ID: yFA4GnzA4

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:55 p.m.

Version: 1

Publication Details
Cremades N;Bueno M;Neira JL;Velázquez-Campoy A;Sancho J,J. Biol. Chem. (2008) Conformational stability of Helicobacter pylori flavodoxin: fit to function at pH 5. PMID:17998211
Additional Information

Study Summary

Number of data points 24
Proteins Flavodoxin ; Flavodoxin
Unique complexes 1
Assays/Quantities/Protocols Experimental Assay: Cm prot_conc:20 microM, pH:9.0, buffers:CHES: 35 mM ; Experimental Assay: m prot_conc:20 microM, pH:9.0, buffers:CHES: 35 mM ; Experimental Assay: dG_H2O prot_conc:20 microM, pH:9.0, buffers:CHES: 35 mM ; Experimental Assay: Cm prot_conc:20 microM, buffers:MOPS: 30 mM, pH:7.0 ; Experimental Assay: m prot_conc:20 microM, buffers:MOPS: 30 mM, pH:7.0 ; Experimental Assay: dG_H2O prot_conc:20 microM, buffers:MOPS: 30 mM, pH:7.0 ; Experimental Assay: Cm prot_conc:20 microM, buffers:Sodium acetate: 16 mM, pH:5.0 ; Experimental Assay: m prot_conc:20 microM, buffers:Sodium acetate: 16 mM, pH:5.0 ; Experimental Assay: dG_H2O prot_conc:20 microM, buffers:Sodium acetate: 16 mM, pH:5.0 ; Experimental Assay: Cm prot_conc:20 microM, pH:2.0, buffers:Sodium phosphate: 23 mM ; Experimental Assay: m prot_conc:20 microM, pH:2.0, buffers:Sodium phosphate: 23 mM ; Experimental Assay: dG_H2O prot_conc:20 microM, pH:2.0, buffers:Sodium phosphate: 23 mM ; Experimental Assay: Cm buffers:CHES: 35 mM, prot_conc:2 microM, pH:9.0 ; Experimental Assay: m buffers:CHES: 35 mM, prot_conc:2 microM, pH:9.0 ; Experimental Assay: dG_H2O buffers:CHES: 35 mM, prot_conc:2 microM, pH:9.0 ; Experimental Assay: Cm buffers:MOPS: 30 mM, prot_conc:2 microM, pH:7.0 ; Experimental Assay: m buffers:MOPS: 30 mM, prot_conc:2 microM, pH:7.0 ; Experimental Assay: dG_H2O buffers:MOPS: 30 mM, prot_conc:2 microM, pH:7.0 ; Experimental Assay: Cm pH:5.0, prot_conc:2 microM, buffers:Sodium acetate: 16 mM ; Experimental Assay: m pH:5.0, prot_conc:2 microM, buffers:Sodium acetate: 16 mM ; Experimental Assay: dG_H2O pH:5.0, prot_conc:2 microM, buffers:Sodium acetate: 16 mM ; Experimental Assay: Cm prot_conc:2 microM, pH:2.0, buffers:Sodium phosphate: 23 mM ; Experimental Assay: m prot_conc:2 microM, pH:2.0, buffers:Sodium phosphate: 23 mM ; Experimental Assay: dG_H2O prot_conc:2 microM, pH:2.0, buffers:Sodium phosphate: 23 mM
Libraries Mutations for sequence MGKIGIFFGTDSGNAEAIAEKISKAIGNAEVVDVAKASKEQFNSFTKVILVAPTAGAGDLQTDWEDFLGTLEASDFANKTIGLVGLGDQDTYSETFAEGIFHIYEKAKAGKVVGQTSTDGYHFEASKAVEGGKFVGLVIDEDNQDDLTDERISKWVEQVKGSFA

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
2BMV 2006-06-22 2.11 Apoflavodoxin from Helicobacter pylori
1FUE 2002-02-06 2.4 FLAVODOXIN FROM HELICOBACTER PYLORI
2W5U 2009-12-22 2.62 Flavodoxin from Helicobacter pylori in complex with the C3 inhibitor

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
97.6 Flavodoxin Q9ZK53 FLAV_HELPJ
100.0 Flavodoxin O25776 FLAV_HELPY