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January 23, 2006
Volume 84, Number 4
p. 11

Structural Biology

New Class Of Topoisomerase

Crystal structure of archaeal enzyme reveals that it is a unique type

Stu Borman

Courtesy of Alfonso Mondragón

One Of A Kind Topoisomerase V structure includes several domains, which are rendered here in different colors. The active site is the ball-and-stick structure near the center.

A topoisomerase enzyme discovered more than a decade ago in an archaeal microorganism turns out to be distinct, both structurally and mechanistically, from any other known topoisomerase. Only four types of topoisomerases were known until now, and the new structure represents a fifth type.

Topoisomerases are essential enzymes in all organisms, from archaea to eukaryotes, including humans. The enzymes ease DNA replication and transcription by introducing transient single- or double-strand breaks into the DNA double helix. They're important targets for drugs, such as the antibiotic fluoroquinolone and the anticancer agent topotecan, and they're widely used to screen for new drugs and as tools in DNA research.

Topoisomerase V was discovered in the archaeal microorganism Methanopyrus kandleri by a group led by Alexei Slesarev, now vice president of R&D at Fidelity Systems, Gaithersburg, Md. (Nature 1993, 364, 735). Alfonso Mondragón, a professor of biochemistry, molecular biology, and cell biology at Northwestern University, and coworkers, including Slesarev, have now determined the first crystal structure of the enzyme, which unexpectedly shows it's in a class of its own (EMBO J., published online Jan. 5,

"It is fascinating to find that archaea have evolved a completely different way of doing a very complicated reaction," Mondragón says. "We still do not know how it works in atomic detail-we are working on that-but it is clear that it will not be in the same way as other topoisomerases."

Type I topoisomerases cut a single strand of double-stranded DNA, whereas type II enzymes cut both strands. Once DNA has been cut by a topoisomerase, the duplex or strands rearrange, changing the degree of coiling. The enzyme subsequently reseals the cut.

Type I enzymes fall into two groups, type IA and IB, depending on whether they form a transient covalent phosphotyrosine bond to the 5' or 3' end of the cut strand. Type II enzymes are of types IIA and IIB, which differ in sequence and three-dimensional structure.

Topoisomerase V cuts single strands and binds to the 3' end of the broken strand, like a IB type, but its active site and overall structure bear little resemblance to those of type IB enzymes.

"This is a very significant finding because the structure is unexpected," says microbiology professor and topoisomerase specialist James J. Champoux of the University of Washington, Seattle. "The novel structure suggests that this archaeal enzyme evolved completely independently of viral, bacterial, and eukaryotic type IB topoisomerases and as such defines a completely new topoisomerase class."

The major question that remains is how topoisomerase V binds DNA, Champoux says. "The authors argue that the protein likely undergoes a major conformation change during DNA binding, and this seems reasonable." It is possible that the conformation at 108 oC, the temperature at which the protein is most active in archaea, is suitable for binding the substrate, whereas the conformation at room temperature, at which the structure was obtained, is not, he says. Champoux notes that a complete description of the mechanism must await a crystal structure of topoisomerase V bound to DNA.

Molecular biotechnologist Vadim V. Demidov of Boston University comments that the new type of topoisomerase could prove useful in DNA sequencing and polymerase chain reaction protocols (when fused with DNA polymerases) or as a source of "helpful construction elements for protein engineering."

Thierry Viard and Kevin D. Corbett, University of California, Berkeley, postdocs who specialize in topoisomerases, believe that topoisomerase V's in vivo functional role warrants further study. They note that the topoisomerase domain analyzed structurally by Mondragón and colleagues is fused in vivo to a DNA repair domain. They comment that the protein might be a repair enzyme with topoisomerase activity rather than a true topoisomerase. Such questions are tough to answer because the enzyme is found in a rare organism that is difficult to study.

"It would be interesting to see if an equivalent enzyme is present in other archaea," Mondragón says, "as this may give us some clues on the origin of this unique protein."

Northwestern University Photo (left), Fidelity Systems Photo (right)

Topo Team The crystal structure of topoisomerase V was determined by (from left in group photo) Mondragón, Bhupesh Taneja, and Asmita Patel and Slesarev (right).

Chemical & Engineering News
ISSN 0009-2347
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