PMID:16339948
| Citation |
Osawa, M and Erickson, HP (2005) Probing the domain structure of FtsZ by random truncation and insertion of GFP. Microbiology (Reading, Engl.) 151:4033-43 |
|---|---|
| Abstract |
Random transposon-mediated mutagenesis has been used to create truncations and insertions of green fluorescent protein (GFP), and Venus-yellow fluorescent protein (YFP), in Escherichia coli FtsZ. Sixteen unique insertions were obtained, and one of them, in the poorly conserved C-terminal spacer, was functional for cell division with the Venus-YFP insert. The insertion of enhanced GFP (eGFP) at this same site was not functional; Venus-YFP was found to be superior to eGFP in other respects too. Testing the constructs for dominant negative effects led to the following general conclusion. The N-terminal domain, aa 1-195, is an independently folding domain that can poison Z-ring function when expressed without a functional C-terminal domain. The effects were weak, requiring expression of the mutant at 3-5 times the level of wild-type FtsZ. The C-terminal domain, aa 195-383, was also independently folding, but had no activity in vivo. The differential activity of the N- and C-terminal domains suggests that FtsZ protofilament assembly is directional, with subunits adding primarily at the bottom of the protofilament. Directional assembly could occur by either a treadmilling or a dynamic instability mechanism. |
| Links |
PubMed Online version:10.1099/mic.0.28219-0 |
| Keywords |
Bacterial Proteins/chemistry; Bacterial Proteins/genetics; Bacterial Proteins/metabolism; Cytoskeletal Proteins/chemistry; Cytoskeletal Proteins/genetics; Cytoskeletal Proteins/metabolism; DNA Transposable Elements; Escherichia coli/chemistry; Escherichia coli/genetics; Escherichia coli Proteins/chemistry; Gene Deletion; Green Fluorescent Proteins/biosynthesis; Green Fluorescent Proteins/chemistry; Green Fluorescent Proteins/genetics; Macromolecular Substances; Mutagenesis, Site-Directed; Recombinant Fusion Proteins/analysis; Recombinant Fusion Proteins/biosynthesis; Recombinant Fusion Proteins/chemistry |
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