Limits...
Structural insight into the DNA-binding mode of the primosomal proteins PriA, PriB, and DnaT.

Huang YH, Huang CY - Biomed Res Int (2014)

Bottom Line: The assembly of the protein complexes within the forked DNA responsible for reloading the replicative DnaB helicase anywhere on the chromosome for genome duplication requires the coordination of transient biomolecular interactions.Over the last decade, investigations on the structure and mechanism of these nucleoproteins have provided considerable insight into primosome assembly.In this review, we summarize and discuss our current knowledge and recent advances on the DNA-binding mode of the primosomal proteins PriA, PriB, and DnaT.

View Article: PubMed Central - PubMed

Affiliation: School of Biomedical Sciences, Chung Shan Medical University, No. 110, Section 1, Chien-Kuo N. Road, Taichung City 40201, Taiwan.

ABSTRACT
Replication restart primosome is a complex dynamic system that is essential for bacterial survival. This system uses various proteins to reinitiate chromosomal DNA replication to maintain genetic integrity after DNA damage. The replication restart primosome in Escherichia coli is composed of PriA helicase, PriB, PriC, DnaT, DnaC, DnaB helicase, and DnaG primase. The assembly of the protein complexes within the forked DNA responsible for reloading the replicative DnaB helicase anywhere on the chromosome for genome duplication requires the coordination of transient biomolecular interactions. Over the last decade, investigations on the structure and mechanism of these nucleoproteins have provided considerable insight into primosome assembly. In this review, we summarize and discuss our current knowledge and recent advances on the DNA-binding mode of the primosomal proteins PriA, PriB, and DnaT.

Show MeSH

Related in: MedlinePlus

Putative dsDNA-binding mode of PriB. The DNA-binding models of PriB are directly constructed by manually superimposing the PriB dimer with DNA-bound crystal structure of RTP (Protein Data Bank entry: 1F4K), HU (Protein Data Bank entry: 1P51), and B-form dsDNA. The hydrophobic (green) and basic residues (blue) of RTP, Lys14, Arg16, Lys51, Arg59, Lys71, Lys74, Lys76, Lys77, Lys81, Lys91, Tyr58, and Tyr88, located on the dsDNA-binding surface, are indicated. The basic residues Arg53, Arg55, Lys56, Arg58, Arg61, Lys64, Lys68, and Arg75 of HU located on the dsDNA-binding surface are also indicated. Considering the known dsDNA-binding sites in PriB, PriB may use the HU-based model to bind dsDNA. Alternatively, PriB may use a similar approach to bind ssDNA and dsDNA because the residues responsible for ssDNA and dsDNA binding are almost overlapped.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4129139&req=5

fig5: Putative dsDNA-binding mode of PriB. The DNA-binding models of PriB are directly constructed by manually superimposing the PriB dimer with DNA-bound crystal structure of RTP (Protein Data Bank entry: 1F4K), HU (Protein Data Bank entry: 1P51), and B-form dsDNA. The hydrophobic (green) and basic residues (blue) of RTP, Lys14, Arg16, Lys51, Arg59, Lys71, Lys74, Lys76, Lys77, Lys81, Lys91, Tyr58, and Tyr88, located on the dsDNA-binding surface, are indicated. The basic residues Arg53, Arg55, Lys56, Arg58, Arg61, Lys64, Lys68, and Arg75 of HU located on the dsDNA-binding surface are also indicated. Considering the known dsDNA-binding sites in PriB, PriB may use the HU-based model to bind dsDNA. Alternatively, PriB may use a similar approach to bind ssDNA and dsDNA because the residues responsible for ssDNA and dsDNA binding are almost overlapped.

Mentions: More than a mere ssDNA-binding protein, PriB can bind both ssDNA and dsDNA with comparable affinity [70]. SSB can also bind dsDNA but with far less affinity than ssDNA [78]. According to the crystal structures of some dimeric proteins complexed with dsDNA found in the Protein Data Bank, PriB binds dsDNA in three possible ways (Figure 5). First, PriB may bind to dsDNA via the replication terminator protein- (RTP-) binding mode (Protein Data Bank entry: 1F4K) [79]. RTP, a dimeric WH protein [80, 81], uses two recognition helices to bind the major grooves of dsDNA. The PriB dimer also has two helices but does not contain any aromatic or positively charged residues as RTP. Thus, PriB binds to dsDNA via the RTP-binding mode that can be ruled out. Second, PriB may bind to dsDNA via the HU-binding mode (Protein Data Bank entry: 1P51) [82, 83]. HU is a dimeric nucleoid-associated protein that mainly uses two β sheets to bind dsDNA. Third, PriB may bind dsDNA in a manner similar to binding ssDNA. The structure-based mutational analysis indicates that the residues in PriB crucial for ssDNA binding are also crucial for dsDNA binding [70]. These residues responsible for ssDNA and dsDNA binding are almost overlapped; thus, PriB may use a similar approach to bind to the phosphate backbone of ssDNA and dsDNA through several positively charged residues. This phenomenon may be the reason for the comparable binding affinities of PriB to ssDNA and dsDNA. We speculate that, during evolution [76], the conserved aromatic and other residues in the L45 loop of the OB fold in SSB are changed into nonconserved and positively charged residues in PriB to more precisely fit the requirement for assembly of the replication restart primosome at the stalled DNA forks.


Structural insight into the DNA-binding mode of the primosomal proteins PriA, PriB, and DnaT.

Huang YH, Huang CY - Biomed Res Int (2014)

Putative dsDNA-binding mode of PriB. The DNA-binding models of PriB are directly constructed by manually superimposing the PriB dimer with DNA-bound crystal structure of RTP (Protein Data Bank entry: 1F4K), HU (Protein Data Bank entry: 1P51), and B-form dsDNA. The hydrophobic (green) and basic residues (blue) of RTP, Lys14, Arg16, Lys51, Arg59, Lys71, Lys74, Lys76, Lys77, Lys81, Lys91, Tyr58, and Tyr88, located on the dsDNA-binding surface, are indicated. The basic residues Arg53, Arg55, Lys56, Arg58, Arg61, Lys64, Lys68, and Arg75 of HU located on the dsDNA-binding surface are also indicated. Considering the known dsDNA-binding sites in PriB, PriB may use the HU-based model to bind dsDNA. Alternatively, PriB may use a similar approach to bind ssDNA and dsDNA because the residues responsible for ssDNA and dsDNA binding are almost overlapped.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4129139&req=5

fig5: Putative dsDNA-binding mode of PriB. The DNA-binding models of PriB are directly constructed by manually superimposing the PriB dimer with DNA-bound crystal structure of RTP (Protein Data Bank entry: 1F4K), HU (Protein Data Bank entry: 1P51), and B-form dsDNA. The hydrophobic (green) and basic residues (blue) of RTP, Lys14, Arg16, Lys51, Arg59, Lys71, Lys74, Lys76, Lys77, Lys81, Lys91, Tyr58, and Tyr88, located on the dsDNA-binding surface, are indicated. The basic residues Arg53, Arg55, Lys56, Arg58, Arg61, Lys64, Lys68, and Arg75 of HU located on the dsDNA-binding surface are also indicated. Considering the known dsDNA-binding sites in PriB, PriB may use the HU-based model to bind dsDNA. Alternatively, PriB may use a similar approach to bind ssDNA and dsDNA because the residues responsible for ssDNA and dsDNA binding are almost overlapped.
Mentions: More than a mere ssDNA-binding protein, PriB can bind both ssDNA and dsDNA with comparable affinity [70]. SSB can also bind dsDNA but with far less affinity than ssDNA [78]. According to the crystal structures of some dimeric proteins complexed with dsDNA found in the Protein Data Bank, PriB binds dsDNA in three possible ways (Figure 5). First, PriB may bind to dsDNA via the replication terminator protein- (RTP-) binding mode (Protein Data Bank entry: 1F4K) [79]. RTP, a dimeric WH protein [80, 81], uses two recognition helices to bind the major grooves of dsDNA. The PriB dimer also has two helices but does not contain any aromatic or positively charged residues as RTP. Thus, PriB binds to dsDNA via the RTP-binding mode that can be ruled out. Second, PriB may bind to dsDNA via the HU-binding mode (Protein Data Bank entry: 1P51) [82, 83]. HU is a dimeric nucleoid-associated protein that mainly uses two β sheets to bind dsDNA. Third, PriB may bind dsDNA in a manner similar to binding ssDNA. The structure-based mutational analysis indicates that the residues in PriB crucial for ssDNA binding are also crucial for dsDNA binding [70]. These residues responsible for ssDNA and dsDNA binding are almost overlapped; thus, PriB may use a similar approach to bind to the phosphate backbone of ssDNA and dsDNA through several positively charged residues. This phenomenon may be the reason for the comparable binding affinities of PriB to ssDNA and dsDNA. We speculate that, during evolution [76], the conserved aromatic and other residues in the L45 loop of the OB fold in SSB are changed into nonconserved and positively charged residues in PriB to more precisely fit the requirement for assembly of the replication restart primosome at the stalled DNA forks.

Bottom Line: The assembly of the protein complexes within the forked DNA responsible for reloading the replicative DnaB helicase anywhere on the chromosome for genome duplication requires the coordination of transient biomolecular interactions.Over the last decade, investigations on the structure and mechanism of these nucleoproteins have provided considerable insight into primosome assembly.In this review, we summarize and discuss our current knowledge and recent advances on the DNA-binding mode of the primosomal proteins PriA, PriB, and DnaT.

View Article: PubMed Central - PubMed

Affiliation: School of Biomedical Sciences, Chung Shan Medical University, No. 110, Section 1, Chien-Kuo N. Road, Taichung City 40201, Taiwan.

ABSTRACT
Replication restart primosome is a complex dynamic system that is essential for bacterial survival. This system uses various proteins to reinitiate chromosomal DNA replication to maintain genetic integrity after DNA damage. The replication restart primosome in Escherichia coli is composed of PriA helicase, PriB, PriC, DnaT, DnaC, DnaB helicase, and DnaG primase. The assembly of the protein complexes within the forked DNA responsible for reloading the replicative DnaB helicase anywhere on the chromosome for genome duplication requires the coordination of transient biomolecular interactions. Over the last decade, investigations on the structure and mechanism of these nucleoproteins have provided considerable insight into primosome assembly. In this review, we summarize and discuss our current knowledge and recent advances on the DNA-binding mode of the primosomal proteins PriA, PriB, and DnaT.

Show MeSH
Related in: MedlinePlus