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Thermal unfolding simulations of bacterial flagellin: insight into its refolding before assembly.

Chng CP, Kitao A - Biophys. J. (2008)

Bottom Line: We observed a similar unfolding order of the domains as reported in experimental thermal denaturation.A recent mutagenesis study on flagellin stability seems to suggest the importance of the folding cores.Using crude size estimates, our data suggests that the chamber might be large enough for either denatured hypervariable-region domains or filament-core domains, but not whole flagellin; this implicates a two-staged refolding process.

View Article: PubMed Central - PubMed

Affiliation: Department of Computational Biology, Graduate School of Frontier Sciences, Institute of Molecular and Cellular Biosciences, The University of Tokyo, Tokyo, Japan.

ABSTRACT
Flagellin is the subunit of the bacterial filament, the micrometer-long propeller of a bacterial flagellum. The protein is believed to undergo unfolding for transport through the channel of the filament and to refold in a chamber at the end of the channel before being assembled into the growing filament. We report a thermal unfolding simulation study of S. typhimurium flagellin in aqueous solution as an attempt to gain atomic-level insight into the refolding process. Each molecule comprises two filament-core domains {D0, D1} and two hypervariable-region domains {D2, D3}. D2 can be separated into subdomains D2a and D2b. We observed a similar unfolding order of the domains as reported in experimental thermal denaturation. D2a and D3 exhibited high thermal stability and contained persistent three-stranded beta-sheets in the denatured state which could serve as folding cores to guide refolding. A recent mutagenesis study on flagellin stability seems to suggest the importance of the folding cores. Using crude size estimates, our data suggests that the chamber might be large enough for either denatured hypervariable-region domains or filament-core domains, but not whole flagellin; this implicates a two-staged refolding process.

Show MeSH
Bacterial flagellin export. A flagellum with a few assembled flagellins near the tip of the filament is shown in this schematic diagram. The assembled or polymeric form of flagellin has been solved (PDB ID: 1UCU), from which the cytoplasmic or monomeric form with disordered termini helices has been obtained from our molecular dynamics simulation in solvent. The conformation of flagellin during transport through the channel is still unknown but is suggested to be highly unfolded due to the narrow channel cross section. Refolding then takes place in the chamber before assembly with the help of HAP2 chaperone. The newly added flagellin shown still has disordered termini helices, with solid circle representing the N-terminal.
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fig1: Bacterial flagellin export. A flagellum with a few assembled flagellins near the tip of the filament is shown in this schematic diagram. The assembled or polymeric form of flagellin has been solved (PDB ID: 1UCU), from which the cytoplasmic or monomeric form with disordered termini helices has been obtained from our molecular dynamics simulation in solvent. The conformation of flagellin during transport through the channel is still unknown but is suggested to be highly unfolded due to the narrow channel cross section. Refolding then takes place in the chamber before assembly with the help of HAP2 chaperone. The newly added flagellin shown still has disordered termini helices, with solid circle representing the N-terminal.

Mentions: The flagellum is the motility device of bacterium, with the long tubelike filament used as a propeller constructed solely of one protein called “flagellin”. Flagellin of Salmonella typhimurium has four domains, with the terminal domain (D0) being highly conserved across bacterial strains (1) as it is involved in forming the inner tube of the filament (2). To reach the growing end of the filament, flagellin synthesized in the cytoplasm has to be exported though a narrow (20 Å diameter) channel that runs through the core of the filament (3). The conformation that flagellin adopts during transport remains a mystery. Fig. 1 shows a schematic diagram of flagellin export from the bacterium cytoplasm to the tip of the growing filament for assembly.


Thermal unfolding simulations of bacterial flagellin: insight into its refolding before assembly.

Chng CP, Kitao A - Biophys. J. (2008)

Bacterial flagellin export. A flagellum with a few assembled flagellins near the tip of the filament is shown in this schematic diagram. The assembled or polymeric form of flagellin has been solved (PDB ID: 1UCU), from which the cytoplasmic or monomeric form with disordered termini helices has been obtained from our molecular dynamics simulation in solvent. The conformation of flagellin during transport through the channel is still unknown but is suggested to be highly unfolded due to the narrow channel cross section. Refolding then takes place in the chamber before assembly with the help of HAP2 chaperone. The newly added flagellin shown still has disordered termini helices, with solid circle representing the N-terminal.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2367190&req=5

fig1: Bacterial flagellin export. A flagellum with a few assembled flagellins near the tip of the filament is shown in this schematic diagram. The assembled or polymeric form of flagellin has been solved (PDB ID: 1UCU), from which the cytoplasmic or monomeric form with disordered termini helices has been obtained from our molecular dynamics simulation in solvent. The conformation of flagellin during transport through the channel is still unknown but is suggested to be highly unfolded due to the narrow channel cross section. Refolding then takes place in the chamber before assembly with the help of HAP2 chaperone. The newly added flagellin shown still has disordered termini helices, with solid circle representing the N-terminal.
Mentions: The flagellum is the motility device of bacterium, with the long tubelike filament used as a propeller constructed solely of one protein called “flagellin”. Flagellin of Salmonella typhimurium has four domains, with the terminal domain (D0) being highly conserved across bacterial strains (1) as it is involved in forming the inner tube of the filament (2). To reach the growing end of the filament, flagellin synthesized in the cytoplasm has to be exported though a narrow (20 Å diameter) channel that runs through the core of the filament (3). The conformation that flagellin adopts during transport remains a mystery. Fig. 1 shows a schematic diagram of flagellin export from the bacterium cytoplasm to the tip of the growing filament for assembly.

Bottom Line: We observed a similar unfolding order of the domains as reported in experimental thermal denaturation.A recent mutagenesis study on flagellin stability seems to suggest the importance of the folding cores.Using crude size estimates, our data suggests that the chamber might be large enough for either denatured hypervariable-region domains or filament-core domains, but not whole flagellin; this implicates a two-staged refolding process.

View Article: PubMed Central - PubMed

Affiliation: Department of Computational Biology, Graduate School of Frontier Sciences, Institute of Molecular and Cellular Biosciences, The University of Tokyo, Tokyo, Japan.

ABSTRACT
Flagellin is the subunit of the bacterial filament, the micrometer-long propeller of a bacterial flagellum. The protein is believed to undergo unfolding for transport through the channel of the filament and to refold in a chamber at the end of the channel before being assembled into the growing filament. We report a thermal unfolding simulation study of S. typhimurium flagellin in aqueous solution as an attempt to gain atomic-level insight into the refolding process. Each molecule comprises two filament-core domains {D0, D1} and two hypervariable-region domains {D2, D3}. D2 can be separated into subdomains D2a and D2b. We observed a similar unfolding order of the domains as reported in experimental thermal denaturation. D2a and D3 exhibited high thermal stability and contained persistent three-stranded beta-sheets in the denatured state which could serve as folding cores to guide refolding. A recent mutagenesis study on flagellin stability seems to suggest the importance of the folding cores. Using crude size estimates, our data suggests that the chamber might be large enough for either denatured hypervariable-region domains or filament-core domains, but not whole flagellin; this implicates a two-staged refolding process.

Show MeSH