PMID:12761391
| Citation |
Mehl, AF, Heskett, LD, Jain, SS and Demeler, B (2003) Insights into dimerization and four-helix bundle formation found by dissection of the dimer interface of the GrpE protein from Escherichia coli. Protein Sci. 12:1205-15 |
|---|---|
| Abstract |
The GrpE heat shock protein from Escherichia coli has a homodimeric structure. The dimer interface encompasses two long alpha-helices at the NH(2)-terminal end from each monomer (forming a "tail"), which lead into a small four-helix bundle from which each monomer contributes two short sequential alpha-helices in an antiparallel topological arrangement. We have created a number of different deletion mutants of GrpE that have portions of the dimer interface to investigate requirements for dimerization and to study four-helix bundle formation. Using chemical crosslinking and analytical ultracentrifugation techniques to probe for multimeric states, we find that a mutant containing only the long alpha-helical tail portion (GrpE1-88) is unable to form a dimer, most likely due to a decrease in alpha-helical content as determined by circular dichroism spectroscopy, thus one reason for a dimeric structure for the GrpE protein is to support the tail region. Mutants containing both of the short alpha-helices (GrpE1-138 and GrpE88-197) are able to form a dimer and presumably the four-helix bundle at the dimer interface. These two mutants have equilibrium constants for the monomer-dimer equilibrium that are very similar to the full-length protein suggesting that the tail region does not contribute significantly to the stability of the dimer. Interestingly, one mutant that contains just one of the short alpha-helices (GrpE1-112) exists as a tetrameric species, which presumably is forming a four-helix bundle structure. A proposed model is discussed for this mutant and its relevance for factors influencing four-helix bundle formation. |
| Links |
PubMed PMC2323885 Online version:10.1110/ps.0300803 |
| Keywords |
Bacterial Proteins/biosynthesis; Bacterial Proteins/chemistry; Bacterial Proteins/genetics; Chromatography, Gel; Circular Dichroism; Cross-Linking Reagents; Dimerization; Electrophoresis, Polyacrylamide Gel; Escherichia coli Proteins/chemistry; Escherichia coli Proteins/genetics; Heat-Shock Proteins/biosynthesis; Heat-Shock Proteins/chemistry; Heat-Shock Proteins/genetics; Mass Spectrometry; Models, Molecular; Monte Carlo Method; Mutation; Sequence Deletion; Ultracentrifugation |
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