The N terminus of MinD contains determinants which affect its dynamic localization and enzymatic activity.
MinD is involved in regulating the proper placement of the cytokinetic machinery in some bacteria, including Neisseria gonorrhoeae and Escherichia coli. Stimulation of the ATPase activity of MinD by MinE has been proposed to induce dynamic, pole-to-pole oscillations of MinD in E. coli. Here, we investigated the effects of deleting or mutating conserved residues within the N terminus of N. gonorrhoeae MinD (MinD(Ng)) on protein dynamism, localization, and interactions with MinD(Ng) and with MinE(Ng). Deletions or mutations were generated in the first five residues of MinD(Ng), and mutant proteins were evaluated by several functional assays. Truncation or mutation of N-terminal residues disrupted MinD(Ng) interactions with itself and with MinE. Although the majority of green fluorescent protein (GFP)-MinD(Ng) mutants could still oscillate from pole to pole in E. coli, the GFP-MinD(Ng) oscillation cycles were significantly faster and were accompanied by increased cytoplasmic localization. Interestingly, in vitro ATPase assays indicated that MinD(Ng) proteins lacking the first three residues or with an I5E substitution possessed higher MinE(Ng)-independent ATPase activities than the wild-type protein. These results indicate that determinants found within the extreme N terminus of MinD(Ng) are implicated in regulating the enzymatic activity and dynamic localization of the protein.
Szeto
Jason
J
Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, Ontario, Canada.
Acharya
Sudeep
S
Eng
Nelson F
NF
Dillon
Jo-Anne R
JA
D000251:Adenosine Triphosphatases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D009344:Neisseria gonorrhoeae / Q000201:enzymology* / Q000235:genetics; D011993:Recombinant Fusion Proteins; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics; D020798:Two-Hybrid System Techniques
D001426:Bacterial Proteins; D011993:Recombinant Fusion Proteins; D000251:Adenosine Triphosphatases; C439050:MinD protein, Neisseria gonorrhoeae