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Literature summary for 5.6.2.1 extracted from

  • Changela, A.; DiGate, R.J.; Mondragon, A.
    Structural studies of E. coli topoisomerase III-DNA complexes reveal a novel type IA topoisomerase-DNA conformational intermediate (2007), J. Mol. Biol., 368, 105-118.
    View publication on PubMedView publication on EuropePMC
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Crystallization (Commentary)

Crystallization (Comment) Organism
expressed in Escherichia coli BL21 cells and purified. Wild type enzyme+ssDNA complex crystallize under the same conditions that produce crystals of the Escherichia coli topoisomerase III (Y328F)+ssDNA complex. The new crystals grow in a different crystal form. In the new crystals, there are two molecules in the asymmetric unit almost related by a 2-fold axis Escherichia coli

Protein Variants

Protein Variants Comment Organism
additional information comparison of wild-type Escherichia coli topoisomerase III with the Escherichia coli topoisomerase III (Y328F) mutant. Conformational differences: the ssDNA is not as deeply buried in the active site of the open complexes as it is in the topoisomerase IIIY328F+ ssDNA complex. Enzyme–DNA interactions along the binding groove in the fully bound wild-type-DNA complex are analogous to those seen in the topoisomerase IIIY328F-DNA structure. Conformation resembles the one observed previously with a DNA-bound, catalytically inactive mutant of topoisomerase III where DNA binding realigns catalytic residues to form a functional active site Escherichia coli

Organism

Organism UniProt Comment Textmining
Escherichia coli P14294
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-

Purification (Commentary)

Purification (Comment) Organism
-
 Escherichia coli

Reaction

Reaction Comment Organism Reaction ID
ATP-independent breakage of single-stranded DNA, followed by passage and rejoining cleaves single-stranded DNA (ssDNA) via a 5' phosphotyrosine intermediate Escherichia coli
a

Synonyms

Synonyms Comment Organism
DNA topoisomerase III belongs to the type IA family of DNA topoisomerases Escherichia coli

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
additional information
-
 When glycerol is used as the cryoprotectant (Form II) at a pH at or below 7, enzyme molecules in the asymmetric unit are found in different conformations, with each one deviating from the apo-form of Escherichia coli topoisomerase III. Crystals soaked in 25% glycerol and at pH 8.0 result in a third structure revealing one molecule in the open conformation with the ssDNA fully binding and the other molecule in an intermediate conformation, where the enzyme is mostly closed, the ssDNA is also in an intermediate conformation between the closed and open forms, and a helix forming part of the groove has shifted Escherichia coli

pH Range

pH Minimum pH Maximum Comment Organism
5.5 8 solved crystal structures of wild-type Escherichia coli topoisomerase III bind to an eight-base ssDNA molecule in three different pH environments. A low pH of the crystallization environment (pH 5.5) inhibits the wild-type enzyme from cleaving its substrate. At pH 7.0, the overall conformation of each molecule in the asymmetric unit remains the same as that observed at pH 5.5. In the pH 8 structure, one of the molecules is in the open conformation and the other in the intermediate conformation, but no cleavage is observed. In all, there are seven different closed structures: one of the molecules in the asymmetric unit in each of the five different conditions (Form I pH 5.5 and 7.0 and Form II pH 5.5, 7.0, and 8.0), and two additional closed complexes in the glucose or Form I crystals. At a pH below 8, crystals cryo-protected with glycerol show one monomer in the closed conformation and one in the open conformation Escherichia coli