Mapping functional domains responsible for NA association and dynamic movement n ParA1 of Streptomyces coelicolor
Date Issued
2009
Date
2009
Author(s)
Wu, Kuan-Yi
Abstract
Chromosome segregation during sporulation in Streptomyces coelicolor A(3)2 is mediated by the Par system. The Par system is consisted of two protein components, ParA1 and ParB, and a centromere-like site, parS. ParA1 belongs to the MinD/ParA subgroup of the Walker-type ATPase family that is characterized by its cytoskeleton properties. Our goal is to gain better understanding on how the Par system may couple its dynamic cytoskeleton property to chromosome segregation. We show here that when expressed in Escherichia coli cells, Yfp tagged ParA1 in the absence of ParB was capable of associating with nucleoids and underwent stochastic movement over the nucleoids. On the other hand, the N-terminal truncated ParA1 preserved its ability to undergo stochastic movement though no longer associated with nucleoids. Both the full-length and N-terminal truncated ParA1 proteins were capable of forming long-range helical cables in E. coli that was independent of actin-like MreB and the Min system. Thus the dynamic movement of ParA1 is independent of ParB and can be separated from nucleoid association. Furthermore, ParA1’s ability to associate with nucleoids was impaired when mutations (R31E, K39E, G40V, K44E, D154A) were introduced into the ATP-binding pocket, suggesting that ATP binding to ParA1 is required for association with nucleoids. Using bacterial two-hybrid assays, ParA1-ParB interaction was enhanced when ParA1 harbored a mutation in Asp68, a residue that is predicted to be involved in ATP hydrolysis. This result indicates that ATP-bound ParA1 may stabilize its interaction with ParB. We also substitute the miniF Par system, SopABC, with Scoe Par system and found that Scoe Par system play active roles in plasmid DNA partition. In summary, we have characterized the functional domains of ParA1 of S. coelicolor A(3)2 that are fundamental for its cytoskeleton properties: (1) the N-terminal domain together with the ATP-binding pocket are responsible for targeting ParA1 to the bacterial nucleoids; (2) the ATP-bound ParA1 is likely to confer the ParB-binding conformation, and (3) the N-terminal domain of ParA1 is not required to deliver the dynamic properties of the protein.
Subjects
chromosome segregation
ParA
ParB
parS
cytoskeleton
Walker type ATPase
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