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Mutagenesis study to disrupt electrostatic interactions on the twofold symmetry interface of Escherichia coli bacterioferritin

Research output: Contribution to journalArticle

Yu Zhang, Lijun Wang, Maziar S. Ardejani, Nur Fazlina Aris, Xun Li, Brendan P. Orner, Fei Wang

Original languageEnglish
Pages (from-to)505-512
Number of pages8
JournalJournal of Biochemistry
Volume158
Issue number6
Early online date26 Jun 2015
DOIs
Publication statusPublished - Dec 2015

King's Authors

Abstract

Ferritins and other cage proteins have been utilized as models to understand the fundamentals of protein folding and self-assembly. The bacterioferritin (BFR) from Escherichia coli, a maxi-ferritin made up of 24 subunits, was chosen as the basis for a mutagenesis study to investigate the role of electrostatic intermolecular interactions mediated through charged amino acids. Through structural and computational analyses, three charged amino acids R30, D56 and E60 which involved in an electrostatic interaction network were mutated to the opposite charge. Four mutants, R30D, D56R, E60H and D56R-E60H, were expressed, purified and characterized. All of the mutants fold into α-helical structures. Consistent with the computational prediction, they all show a lowered thermostability; double mutant D56R-E60H was found to be 16°C less stable than the wild type. Except for the mutant E60H, all the other mutations completely shut down the formation of protein cages to favour the dimer state in solution. The mutants, however, retain their ability to form cage-like nanostructures in the dried, surface immobilized conditions of transmission electron microscopy. Our findings confirm that even a single charge-inversion mutation at the 2-fold interface of BFR can affect the quaternary structure of its dimers and their ability to self-assemble into cage structures.

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