The tail of the ParG DNA segregation protein remodels ParF polymers and enhances ATP hydrolysis via an arginine finger-like motif

TY - JOUR

T1 - The tail of the ParG DNA segregation protein remodels ParF polymers and enhances ATP hydrolysis via an arginine finger-like motif

AU - Barilla, Daniela

AU - Carmelo, Emma

AU - Hayes, Finbarr

PY - 2007/2

Y1 - 2007/2

N2 - The ParF protein of plasmid TP228 belongs to the ubiquitous superfamily of ParA ATPases that drive DNA segregation in bacteria. ATF-bound ParF polymerizes into multistranded filaments. The partner protein ParG is dimeric, consisting of C-termini that interweave into a ribbon-helix-helix domain contacting the centromeric DNA and unstructured N-termini. ParG stimulates ATP hydrolysis by ParF approximate to 30-fold. Here, we establish that the mobile tails of ParG are crucial for this enhancement and that arginine R19 within the tail is absolutely required for activation of ParF nucleoticle hydrolysis. R19 is part of an arginine finger-like loop in ParG that is predicted to intercalate into the ParF nucleotide-binding pocket thereby promoting ATP hydrolysis. Significantly, mutations of R19 abrogated DNA segregation in vivo, proving that intracellular stimulation of ATP hydrolysis by ParG is a key regulatory process for partitioning. Furthermore, ParG bundles ParF-ATP filaments as well as promoting nucleotide-independent polymerization. The N-terminal flexible tail is required for both activities, because N-terminal Delta ParG polypepticles are defective in both functions. Strikingly, the critical arginine finger-like residue R19 is dispensable for ParG-mediated remodeling of ParF polymers, revealing that the ParG N-terminal tail possesses two separable activities in the interplay with ParF: a catalytic function during ATP hydrolysis and a mechanical role in modulation of polymerization. We speculate that activation of nucleoticle hydrolysis via an arginine finger loop may be a conserved, regulatory mechanism of ParA family members and their partner proteins, including ParAParB and Soj-SpoOJ that mediate DNA segregation and MinD-MinE that determine septum localization.

AB - The ParF protein of plasmid TP228 belongs to the ubiquitous superfamily of ParA ATPases that drive DNA segregation in bacteria. ATF-bound ParF polymerizes into multistranded filaments. The partner protein ParG is dimeric, consisting of C-termini that interweave into a ribbon-helix-helix domain contacting the centromeric DNA and unstructured N-termini. ParG stimulates ATP hydrolysis by ParF approximate to 30-fold. Here, we establish that the mobile tails of ParG are crucial for this enhancement and that arginine R19 within the tail is absolutely required for activation of ParF nucleoticle hydrolysis. R19 is part of an arginine finger-like loop in ParG that is predicted to intercalate into the ParF nucleotide-binding pocket thereby promoting ATP hydrolysis. Significantly, mutations of R19 abrogated DNA segregation in vivo, proving that intracellular stimulation of ATP hydrolysis by ParG is a key regulatory process for partitioning. Furthermore, ParG bundles ParF-ATP filaments as well as promoting nucleotide-independent polymerization. The N-terminal flexible tail is required for both activities, because N-terminal Delta ParG polypepticles are defective in both functions. Strikingly, the critical arginine finger-like residue R19 is dispensable for ParG-mediated remodeling of ParF polymers, revealing that the ParG N-terminal tail possesses two separable activities in the interplay with ParF: a catalytic function during ATP hydrolysis and a mechanical role in modulation of polymerization. We speculate that activation of nucleoticle hydrolysis via an arginine finger loop may be a conserved, regulatory mechanism of ParA family members and their partner proteins, including ParAParB and Soj-SpoOJ that mediate DNA segregation and MinD-MinE that determine septum localization.

KW - ParA superfamily

KW - polymerization

KW - plasmid partition

KW - ATPase

KW - TOPOLOGICAL SPECIFICITY

KW - ESCHERICHIA-COLI

KW - P1 PARA

KW - PLASMID SEGREGATION

KW - BACTERIAL PLASMIDS

KW - STRUCTURAL BASIS

KW - CELL-DIVISION

KW - BINDING SITE

KW - MINE

KW - PARTITION

U2 - 10.1073/pnas.0607216104

DO - 10.1073/pnas.0607216104

M3 - Article

SN - 0027-8424

VL - 104

SP - 1811

EP - 1816

JO - Proceedings of the National Academy of Sciences of the United States of America

JF - Proceedings of the National Academy of Sciences of the United States of America

IS - 6

ER -