PMID:20179345
| Citation |
Terakado, K, Kodan, A, Nakano, H, Kimura, Y, Ueda, K, Nakatsu, T and Kato, H (2010) Deleting two C-terminal alpha-helices is effective to crystallize the bacterial ABC transporter Escherichia coli MsbA complexed with AMP-PNP. Acta Crystallogr. D Biol. Crystallogr. 66:319-23 |
|---|---|
| Abstract |
An MsbA deletion mutant DeltaC21 that lacks the two C-terminal alpha-helices was expressed in Escherichia coli strain C41 and purified by metal-affinity and gel-filtration chromatography. Purified DeltaC21 retained 26% of the activity of the wild-type ATPase and had a similar binding affinity to fluorescent nucleotide derivatives. Although crystals of wild-type MsbA complexed with adenosine 5'-(beta,gamma-imido)triphosphate could not be obtained, crystals of DeltaC21 that diffracted to 4.5 A resolution were obtained. The preliminary DeltaC21 structure had the outward-facing conformation, in contrast to the previously reported E. coli MsbA structure. This result suggests that deletion of the C-terminal alpha-helices may play a role in facilitating the outward-facing nucleotide-bound crystal structure of EcMsbA. |
| Links |
PubMed Online version:10.1107/S0907444909055504 |
| Keywords |
ATP-Binding Cassette Transporters/chemistry; ATP-Binding Cassette Transporters/metabolism; Adenylyl Imidodiphosphate/chemistry; Adenylyl Imidodiphosphate/metabolism; Amino Acid Sequence; Bacterial Proteins/chemistry; Bacterial Proteins/metabolism; Crystallography, X-Ray; Escherichia coli/chemistry; Escherichia coli/metabolism; Models, Molecular; Molecular Sequence Data; Protein Binding; Protein Structure, Quaternary; Protein Structure, Secondary; Protein Structure, Tertiary; Sequence Alignment |
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