Allosteric interactions between the nucleotide-binding sites and the ssDNA-binding site in the PriA helicase-ssDNA complex. 3

Aaron L. Lucius, Maria J. Jezewska, Wlodzimierz Bujalowski

Research output: Contribution to journalArticle

13 Citations (Scopus)

Abstract

Allosteric interactions between the strong and weak nucleotide-binding sites and the total and proper single-stranded (ss)DNA-binding sites of the Escherichia coli PriA helicase have been analyzed using the fluorescence titration technique. Binding of the DNA exclusively to the proper DNA-binding site of the helicase, profoundly affects the intrinsic affinities of both nucleotide-binding sites, indicating a direct communication between the nucleotide-binding sites and the proper DNA-binding site. The communication involves conformational changes of the entire protein molecule. Nevertheless, the bound DNA differently affects the structures of the strong and weak nucleotide-binding sites. While the polarity of the strong site is moderately diminished, the polarity of the weak site is dramatically increased, indicating an intimate involvement of the weak site in controlling the helicase interactions with the DNA. The strong site does not directly control the DNA affinity of the enzyme. Only when the helicase has both nucleotide-binding sites saturated with ADP but not with ATP analogues does the enzyme have an increased affinity for the ssDNA, indicating that the control of ssDNA affinity involves a coordinated action of both nucleotide-binding sites and depends upon the phosphate group of the bound cofactor. A dramatic increase of the DNA affinity, when the DNA encompasses the total DNA-binding site of the enzyme, with both nucleotide-binding sites saturated with ADP or NDP, indicates that an additional area of the protein within the total DNA-binding site becomes engaged in interactions with the DNA. The significance of these results for the enzyme activities in the DNA unwinding and recognition is discussed.

Original languageEnglish (US)
Pages (from-to)7237-7255
Number of pages19
JournalBiochemistry
Volume45
Issue number23
DOIs
StatePublished - Jun 13 2006

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Nucleotides
Binding Sites
DNA
Enzymes
Adenosine Diphosphate
Single-Stranded DNA
Communication
Enzyme activity
Titration
Escherichia coli
Proteins
Adenosine Triphosphate
Fluorescence
Phosphates
Molecules

ASJC Scopus subject areas

  • Biochemistry

Cite this

Allosteric interactions between the nucleotide-binding sites and the ssDNA-binding site in the PriA helicase-ssDNA complex. 3. / Lucius, Aaron L.; Jezewska, Maria J.; Bujalowski, Wlodzimierz.

In: Biochemistry, Vol. 45, No. 23, 13.06.2006, p. 7237-7255.

Research output: Contribution to journalArticle

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