Hda Monomerization by ADP Binding Promotes Replicase Clamp-mediated DnaA-ATP Hydrolysis

Abstract

ATP-DnaA is the initiator of chromosomal replication in Escherichia coli, and the activity of DnaA is regulated by the regulatory inactivation of the DnaA (RIDA) system. In this system, the Hda protein promotes DnaA-ATP hydrolysis to produce inactive ADP-DnaA in a mechanism that is mediated by the DNA-loaded form of the replicase sliding clamp. In this study, we first revealed that hda translation uses an unusual initiation codon, CUG, located downstream of the annotated initiation codon. The CUG initiation codon could be used for restricting the Hda level, as this initiation codon has a low translation efficiency, and the cellular Hda level is only ∼100 molecules per cell. Hda translated using the correct reading frame was purified and found to have a high RIDA activity in vitro. Moreover, we found that Hda has a high affinity for ADP but not for other nucleotides, including ATP. ADP-Hda was active in the RIDA system in vitro and stable in a monomeric state, whereas apo-Hda formed inactive homomultimers. Both ADP-Hda and apo-Hda could form complexes with the DNA-loaded clamp; however, only ADP-Hda-DNA-clamp complexes were highly functional in the following interaction with DnaA. Formation of ADP-Hda was also observed in vivo, and mutant analysis suggested that ADP binding is crucial for cellular Hda activity. Thus, we propose that ADP is a crucial Hda ligand that promotes the activated conformation of the protein. ADP-dependent monomerization might enable the arginine finger of the Hda AAA+ domain to be accessible to ATP bound to the DnaA AAA+ domain. ATP-DnaA is the initiator of chromosomal replication in Escherichia coli, and the activity of DnaA is regulated by the regulatory inactivation of the DnaA (RIDA) system. In this system, the Hda protein promotes DnaA-ATP hydrolysis to produce inactive ADP-DnaA in a mechanism that is mediated by the DNA-loaded form of the replicase sliding clamp. In this study, we first revealed that hda translation uses an unusual initiation codon, CUG, located downstream of the annotated initiation codon. The CUG initiation codon could be used for restricting the Hda level, as this initiation codon has a low translation efficiency, and the cellular Hda level is only ∼100 molecules per cell. Hda translated using the correct reading frame was purified and found to have a high RIDA activity in vitro. Moreover, we found that Hda has a high affinity for ADP but not for other nucleotides, including ATP. ADP-Hda was active in the RIDA system in vitro and stable in a monomeric state, whereas apo-Hda formed inactive homomultimers. Both ADP-Hda and apo-Hda could form complexes with the DNA-loaded clamp; however, only ADP-Hda-DNA-clamp complexes were highly functional in the following interaction with DnaA. Formation of ADP-Hda was also observed in vivo, and mutant analysis suggested that ADP binding is crucial for cellular Hda activity. Thus, we propose that ADP is a crucial Hda ligand that promotes the activated conformation of the protein. ADP-dependent monomerization might enable the arginine finger of the Hda AAA+ domain to be accessible to ATP bound to the DnaA AAA+ domain. The initiation of chromosomal replication is strictly regulated during the cell cycle. In Escherichia coli, a crucial target for this regulation is the formation of an active initiation complex, including the ATP-bound DnaA protein (ATP-DnaA) and the chromosomal replication origin, oriC (1Messer W. FEMS Microbiol. Rev. 2002; 26: 355-374PubMed Google Scholar, 2Kaguni J.M. Annu. Rev. Microbiol. 2006; 60: 351-375Crossref PubMed Scopus (165) Google Scholar). The DiaA protein directly stimulates formation of this complex, which leads to the unwinding of duplex DNA within the oriC (3Ishida T. Akimitsu N. Kashioka T. Hatano M. Kubota T. Ogata Y. Sekimizu K. Katayama T. J. Biol. Chem. 2004; 279: 45546-45555Abstract Full Text Full Text PDF PubMed Scopus (94) Google Scholar, 4Keyamura K. Fujikawa N. Ishida T. Ozaki S. Su'etsugu M. Fujimitsu K. Kagawa W. Yokoyama S. Kurumizaka H. Katayama T. Genes Dev. 2007; 21: 2083-2099Crossref PubMed Scopus (102) Google Scholar). DnaB helicase then expands the unwound region to allow the loading of DnaG primase and DNA polymerase (pol) 5The abbreviations used are: polDNA polymeraseRIDAregulatory inactivation of DnaASDShine-DalgarnoATPγSadenosine 5′-O-(thiotriphosphate)FrfractionntHdanontagged form of Hda III holoenzyme (5O'Donnell M. J. Biol. Chem. 2006; 281: 10653-10656Abstract Full Text Full Text PDF PubMed Scopus (104) Google Scholar). The III holoenzyme of the III and The a as a and is the DNA by the to the III DNA during DNA of the the III and the sliding the DNA (5O'Donnell M. J. Biol. Chem. 2006; 281: 10653-10656Abstract Full Text Full Text PDF PubMed Scopus (104) Google Scholar). DNA polymerase regulatory inactivation of DnaA form of Hda that J.M. Annu. Rev. Microbiol. 2006; 60: 351-375Crossref PubMed Scopus (165) Google Scholar, T. Microbiol. PubMed Scopus Google Scholar, J. W. FEMS Microbiol. Rev. 2007; PubMed Scopus Google Scholar). binding of to oriC initiation formation M. N. Full Text PDF PubMed Scopus Google Scholar, S. S. M. K. N. Full Text PDF PubMed Scopus Google Scholar, 2006; Full Text Full Text PDF PubMed Scopus Google Scholar). The oriC region that for by the a oriC that is the target of In the system, is a of DnaA molecules that accessible to the The is located the oriC and to a of DnaA molecules Ozaki T. S. T. Genes Dev. PubMed Scopus Google Scholar). of DnaA molecules is by of the following The system the regulatory inactivation of DnaA (RIDA) and promotes the hydrolysis of which inactive ADP-DnaA J. Katayama T. J. PubMed Scopus Google Scholar, T. Kubota T. K. Sekimizu K. Full Text Full Text PDF PubMed Scopus Google Scholar). RIDA the Hda protein and the DNA-loaded form of the clamp. The of the DNA-loaded inactivation of DnaA during the replication of the The cellular ATP-DnaA level to the initiation of replication and chromosomal replication J. Katayama T. J. PubMed Scopus Google Scholar, K. S. Sekimizu K. Katayama T. J. PubMed Scopus Google Scholar). a mutant of DnaA of ATP-DnaA and initiation J. Katayama T. J. PubMed Scopus Google Scholar, S. Fujimitsu K. Sekimizu K. T. T. Katayama T. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar, M. H. K. Kubota T. M. Katayama T. Microbiol. PubMed Scopus Google Scholar, T. S. PubMed Scopus Google Scholar). The of replication and initiation regulation to be and as a of initiation by a with DnaA and the clamp; however, this protein with the of Hda J. S. 2002; PubMed Scopus Google Scholar, M. Y. N. S. FEMS Microbiol. PubMed Scopus Google Scholar, M. M. S. T. N. S. 2006; PubMed Scopus Google Scholar). The sliding cell with the replication Biol. 2006; PubMed Scopus Google Scholar, H. N. Y. M. T. K. M. T. J. 2006; PubMed Scopus Google Scholar, T. W. J. Biol. Chem. 2006; 281: Full Text Full Text PDF PubMed Scopus Google Scholar). interaction promotes of which is with the of In a RIDA Hda promotes DnaA-ATP hydrolysis in the of the DNA-loaded M. M. Kubota T. Y. Katayama T. Genes 2004; PubMed Scopus Google Scholar, M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). the binding to Hda region that the M. M. Kubota T. Y. Katayama T. Genes 2004; PubMed Scopus Google Scholar, M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, K. S. PubMed Scopus Google Scholar, M. K. J. 2004; PubMed Scopus Google Scholar). the Hda an AAA+ domain that is with the DnaA AAA+ domain J. Katayama T. J. PubMed Scopus Google Scholar). domain and that in the AAA+ PubMed Google Scholar). of AAA+ is the formation of in which ATP binding and hydrolysis the and a In this a arginine an arginine finger a crucial in the of ATP bound to the J. M. Rev. Biol. 2002; PubMed Scopus Google Scholar, T. J. Biol. 2004; PubMed Scopus Google Scholar). and of and the DNA-loaded the following is M. M. Kubota T. Y. Katayama T. Genes 2004; PubMed Scopus Google Scholar, M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The Hda arginine finger directly promotes ATP hydrolysis interaction the AAA+ of DnaA and The DNA-loaded this interaction as a which the DnaA domain with the complex, and the DnaA domain the duplex DNA the clamp. we found that Hda to ADP but not ATP in vitro and in formed a that is to of in the of ADP binding of the Hda to and activated Hda in the RIDA system in vitro. in vitro and in using the of Hda suggested that ADP binding is crucial for Hda in ADP-Hda but not apo-Hda formed a functional with ATP-DnaA and the DNA-loaded clamp. The Hda arginine finger could be the of apo-Hda and ADP-dependent monomerization might the arginine finger the interaction with and of Hda for the and were M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). was by and of a using as a and the and of S. Fujimitsu K. Sekimizu K. T. T. Katayama T. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google were in and the of the was and were for an of the of of using was using the as that for a Hda M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). was by a and by an using a protein and of the region was by using as a and the and and the and of in were in the of the was and were for the following were of a cell and were by the as that for a Hda M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google the and was using a of a with and of was for in the of were by in and a with a of of and an was purified an of cell The cell was and protein were by the as that for Hda M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The were in and a using the as that for for mutant were using the and of was using an cell and the as and of for was by and of a using as a and the and and were by the as that for Hda M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google was a with a of and including the were were as M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google that was the protein of the ADP were for in and the of were by and of the that was the K. Full Text PDF PubMed Scopus Google Scholar). of a ATP was a with and and with a of to of ATP and ADP was The of ATP was in and a with ATP was with and a RIDA RIDA system was used as M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). In the first the and DNA were for in of and ATP and The DNA-loaded was in a by a using with In the the DNA-loaded molecules of the per DNA was for in DnaA and Hda in the of ATP ADP bound to DnaA were a and using and a In of Hda in DnaA-ATP an region and was by using DNA and the and the and of in The was the of in was using the hda of and J. Katayama T. J. PubMed Scopus Google were in and but and to an of as T. Kubota T. K. Sekimizu K. Full Text Full Text PDF PubMed Scopus Google Scholar). DnaA was by and the bound to the DnaA were using by In of and cell were using the for the in analysis of DnaA were in and of of the was and for in the were by a in for in the of in and and in were by and for with in the of The were and in and the were in in the were using and a of and of by for of DnaA and Hda with the a of the DNA-loaded the and DNA were for in ATP and of The was a with a of the DNA-loaded in the molecules of the per DNA The DNA-loaded was for in by a in the of ATP-DnaA the and as was only the was and the was a with and the DNA and bound were the by a for of DnaA and Hda with the DNA-loaded was using the as for the the was for in the of in and The were for in the of the DNA-loaded and by for in the of by and in The were in and The hda CUG of the we an the annotated region of the hda M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). In the of we that the of Hda by protein and the and was that of Hda the chromosomal could not this the however, in this study, we the and were to in the hda region to as the annotated initiation codon M. J. J. Y. PubMed Scopus Google and the were with and the M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). the was to we the of Hda by the chromosomal and which the chromosomal region of the hda The of Hda was to that of the chromosomal Hda and that of In Hda was in of Hda cell which is by of oriC and J. Katayama T. J. PubMed Scopus Google Scholar). The to the in a of DnaA and oriC T. Microbiol. Biol. Rev. PubMed Scopus Google Scholar). the of the we a a form of Hda using purified revealed that the was which that the initiation codon used in is the CUG that is located downstream the annotated initiation codon of the CUG to Hda CUG for The of the Hda translated the CUG was to be this that the of the Hda was the of which is with the of the analysis the that the CUG as the initiation codon of The in is in which might the of this protein in and might in a in the and the RIDA of the activity of in the RIDA system, we using of the was in a cell and was used for by The of the was analysis of revealed and with a of the of the to the of and the to the of of the activity of for DnaA-ATP hydrolysis in a system M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). the of Hda in a we first the DNA-loaded form of the clamp. was a using the the and form of DNA as M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The DNA-loaded was then to a and ATP. DnaA-ATP hydrolysis the DNA-loaded The activity of was that of is an an this protein was used and was M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The could be to Hda activity in was purified by a to ADP to AAA+ an activity for the binding activity of using a a binding was for binding for ADP was The and the binding were a in and per Hda binding of ADP and is Hda was formed and in the of of was observed in a that the of Hda ADP which might be to a in the binding of Hda for of was by a using The of was that of were with the of ADP of ADP binding was observed in the of which that the affinity of Hda for ATP is that of used a purified ATP with a ADP level of but we were not to the that was to an level of that the of ATP is only that of ADP J. M. Escherichia and and for the high of Hda in binding is with the that in Hda molecules with ADP of in a ADP the of Hda the of ADP Hda activity in DnaA-ATP we the RIDA The DNA-loaded were and in Hda and ATP-DnaA in the of was Hda activity was ADP the of ADP using Hda and that were and and The activity of the was to that of and and was not in the of ADP In the activity of the was that of the in the of ADP Moreover, the of ADP the activity of the by an that the ADP form of Hda is monomeric and active in the DnaA-ATP whereas the the inactive is with the that the form was in and was active in DnaA-ATP is that the in were activated by in a in the activity of the was in the of which might be to the of ADP during the The activity of the in the of ADP was that of the and which might be to the that the ADP to Hda a ADP the Hda that the Hda of whereas the were the ADP form and active in is with the that ADP-Hda complexes stable and also with the that ADP-Hda formed in was during the that was this we apo-Hda using the were was in the of to ADP that was with the Hda this of the Hda molecules formed which were in including the protein was a with ADP and Hda was in a to as and was we the III using Hda was as a a to In the first the for protein not be in the of an form of Hda The of the to but the might of of Hda molecules Hda in the is the III was inactive and in an ADP-dependent III was in the of ADP and by the of Hda was as a in the of the with the that apo-Hda was formed by with and in in ADP-dependent DnaA-ATP hydrolysis were the in the were used not ADP binding and the ADP-dependent RIDA of III were in with of III ADP binding activity a level to that of binding of ADP was observed but with the using and In ADP binding was ADP-Hda was stable the low Hda an inactive conformation that of ADP in to of ADP bound a ADP binding activity which might to a of Hda in The for ADP was to that of III was in the of ADP and was with ATP-DnaA and the DNA-loaded the activity was a level to that of that the and not Hda and that III is in RIDA activity to the that apo-Hda that inactive in RIDA and that ADP binding Hda by the to ADP the of the of we that was an the region purified using a to that of and The ADP binding activity of was that of that ADP binding is by the was by the that but not and and The the activity for DnaA-ATP hydrolysis a level with that of III in the of ADP of Hda to bound to Hda in the were in and for in the of was using a and the were by The of a to ADP with and is with the in vitro that Hda to ADP ATP and the other The Hda in was that in the as by analysis M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). of the was to be and the of the ADP per Hda was to be In the used for Hda was that Hda with ADP is in of Hda in ADP in the in of ADP binding to we Hda in the and Hda Hda and Hda of and Hda were as to using the The mutant Hda formed whereas of were in a The and mutant in the were with and in the was by In the the of was the of the mutant in was by of the to with and of protein as in the binding activity and the binding activity not that the and mutant formed not The purified was active in DnaA-ATP hydrolysis and ADP binding with of purified a The revealed that Hda was in ADP binding In with DnaA-ATP hydrolysis activity of Hda in the of ADP was and The binding activity was in Hda the that of this mutant protein is In to Hda Hda and Hda in ADP binding and DnaA-ATP hydrolysis The ADP binding activity of Hda was that of the protein with the activity of Hda in DnaA-ATP hydrolysis was that of the protein and In the ADP binding activity of Hda was but activity was only that of Hda The DnaA-ATP hydrolysis activity of Hda was but a activity was a level to that of Hda a the ADP binding and RIDA activity of for ADP to Hda in ATP-DnaA level is to of the in an in a J. Katayama T. J. PubMed Scopus Google Scholar, K. S. Sekimizu K. Katayama T. J. PubMed Scopus Google Scholar). hda leads to of the ATP-DnaA level to ADP is for the of Hda in vivo, we the ATP-DnaA in the within the form of mutant hda were a low was used to of the not of the In with the cellular ATP-DnaA level was to in a that was the cellular and and In the ATP-DnaA level was to and that this Hda mutant was in DnaA-ATP hydrolysis in is with the in vitro in ADP binding and DnaA-ATP hydrolysis in this mutant Hda an ATP-DnaA level that was to that in Hda and and is that the of Hda and Hda that were observed in vitro in in The Hda in the the hda were with that in a and of ADP J. M. Escherichia and and for Scholar). The cellular of was with that of and Thus, the ATP-DnaA level in could not be to a in cellular Hda the that Hda ADP for cellular activity. of ADP Formation of Hda with the DNA-loaded the mechanism by which ADP promotes Hda activity in the RIDA system. we used a to the of ADP-Hda and apo-Hda in a formation with the DNA-loaded clamp. In this we used and Hda that were in the in was formed by in the of In a and were in a The of per was that of this is to the that whereas is monomeric The complexes of and the DNA-loaded were active in the DnaA-ATP whereas of and the DNA-loaded were inactive not the of we a analysis using Hda Hda binding activity a level to that of Hda in the of ADP The of but not Hda was Hda was with is with the that Hda is inactive in ADP binding and a using the DNA-loaded and In this we used of binding of to were for in the of in the ADP of the and DNA were the and and that ADP not the affinity of Hda for the DNA-loaded clamp. ADP-dependent Formation of Hda with DnaA by the DNA-loaded of the RIDA we have that DnaA-ATP hydrolysis the formation of a in which ATP-DnaA with Hda in a the DNA-loaded clamp. the of ADP binding to Hda during we first the of the the DNA-loaded and were and ATP-DnaA was by DnaA was in a that was the DNA-loaded and was in the DnaA The of were of were observed for the of and and to the formation of a of DnaA and Hda was the DNA was used and which is to a DNA binding activity of that ADP stimulates Hda activity in with DnaA. we a using ATP-DnaA was in in the of the DNA-loaded by the of using but not the of DnaA with the DnaA was in a the DNA-loaded but not the with the that RIDA the DNA-loaded clamp. The of the and Hda were not by ADP binding to that ADP binding of Hda stimulates stable interaction with ATP-DnaA Hda is with the DNA-loaded clamp. In this study, we that Hda is with the unusual of binding to ADP but not and that ADP is the crucial of ADP-Hda to is mediated by of apo-Hda ADP-Hda In we have that ADP-Hda be formed in and that an Hda mutant in ADP binding was inactive in of the cellular ATP-DnaA Moreover, for the first we have in vitro that and the DNA-loaded form a apo-Hda form a with the DNA-loaded clamp. In but not is the active form for the ATP-DnaA interaction with the Thus, we have revealed ADP-dependent of Hda and Hda might be to as a regulatory for the RIDA system in the monomerization of inactive of the in this is that the hda initiation codon is CUG In coli, the of and as initiation is and J. Biol. PubMed Scopus Google Scholar). In has to CUG as an initiation codon in The CUG initiation codon translation using the that the activity of CUG in initiation is that of Microbiol. 21: PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar). In and to CUG as an initiation codon. the of and the protein of translated the CUG initiation codon S. H. Biol. PubMed Scopus Google Scholar, 2006; PubMed Scopus Google Scholar). In the CUG initiation codon is to be used for of the have that the cellular of the Hda is only ∼100 per cell M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The CUG initiation codon might be used to the low level of cellular Hda molecules as of Hda is to cell J. Katayama T. J. PubMed Scopus Google Scholar, T. N. N. J. W. J. PubMed Scopus Google Scholar). have used the that were a annotated region a affinity for ADP that the region was in ADP binding The of AAA+ has revealed that the of the AAA+ domain is located a to the Rev. Biol. PubMed Scopus Google Scholar). The region is in H. Su'etsugu M. Katayama T. J. Biol. 2006; PubMed Scopus Google which might with the in the of ADP the DnaA-ATP hydrolysis activity of ADP not Thus, the affinity for ADP-dependent of Hda was in In in vitro RIDA ATP was J. Katayama T. J. PubMed Scopus Google Scholar, T. Kubota T. K. Sekimizu K. Full Text Full Text PDF PubMed Scopus Google Scholar, S. Fujimitsu K. Sekimizu K. T. T. Katayama T. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar, M. H. K. Kubota T. M. Katayama T. Microbiol. PubMed Scopus Google Scholar, M. M. Kubota T. Y. Katayama T. Genes 2004; PubMed Scopus Google Scholar, M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The ATP a ADP that was by of ATP. Thus, the RIDA which was high to ADP binding of Hda formed in M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google and The of the might be to the In the of and the of formed complexes as also in M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). was for the stable of activity in M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). In the of and the of Hda formed of the not other AAA+ Hda only ADP but not ATP and and could be with the within the Hda in that Hda the of the The within with the and T. Genes PubMed Scopus Google Scholar). The of Hda is and might with ATP the binding AAA+ the of III the as as affinity for ATP and ADP M. J. Full Text Full Text PDF PubMed Scopus Google Scholar). The an arginine to the AAA+ arginine finger and this for ATP that is bound the in the complex, to ATP hydrolysis M. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The arginine finger of Hda is crucial for DnaA-ATP hydrolysis M. T. Ishida T. H. Katayama T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). In this the and Hda might a that not ATP and the arginine finger with ATP bound to protein ATP is not for the of ADP binding of Hda was for the stable monomeric In of AAA+ the a of the ADP binding might a a in this in of the interaction Hda In AAA+ ADP the formation of the ADP form of a and an of whereas ATP M. J. PubMed Scopus Google Scholar, PubMed Scopus Google Scholar). is by but not ADP H. Biol. PubMed Scopus Google Scholar). In apo-Hda is inactive for DnaA-ATP whereas ADP-Hda is active and of this is to AAA+ that to form including for AAA+ protein the arginine finger of apo-Hda might be in the of the in of the interaction with DnaA. ADP the Hda the arginine finger the protein interaction with DnaA. analysis suggested that ADP binding is crucial for Hda activity in vitro and in and is a that Hda activity might be by and of ADP in the cellular ADP has to be J. M. Escherichia and and for is to the of a ADP binding to In be to a ADP binding of Hda is In a of replication initiation of the S. J. Biol. PubMed Scopus Google Scholar). RIDA might be in the of the DNA-loaded for an in the ATP-DnaA is that a the ADP-Hda a regulatory by RIDA in a be to for a and to the cellular of the cell in to and Yokoyama for analysis and with