title: Insights from the Structure of Mycobacterium tuberculosis Topoisomerase I with a Novel Protein Fold.
abstract: The DNA topoisomerase I enzyme of Mycobacterium tuberculosis (MtTOP1) is essential for the viability of the organism and survival in a murine model. This topoisomerase is being pursued as a novel target for the discovery of new therapeutic agents for the treatment of drug-resistant tuberculosis. In this study, we succeeded in obtaining a structure of MtTOP1 by first predicting that the C-terminal region of MtTOP1 contains four repeated domains that do not involve the Zn-binding tetracysteine motifs seen in the C-terminal domains of Escherichia coli topoisomerase I. A construct (amino acids A2-T704), MtTOP1-704t, that includes the N-terminal domains (D1-D4) and the first predicted C-terminal domain (D5) of MtTOP1 was expressed and found to retain DNA cleavage-religation activity and catalyze single-stranded DNA catenation. MtTOP1-704t was crystallized, and a structure of 2.52Å resolution limit was obtained. The structure of the MtTOP1 N-terminal domains has features that have not been observed in other previously available bacterial topoisomerase I crystal structures. The first C-terminal domain D5 forms a novel protein fold of a four-stranded antiparallel β-sheet stabilized by a crossing-over α-helix. Since there is only one type IA topoisomerase present in Mycobacteriaceae and related Actinobacteria, this subfamily of type IA topoisomerase may be required for multiple functions in DNA replication, transcription, recombination, and repair. The unique structural features observed for MtTOP1 may allow these topoisomerase I enzymes to carry out physiological functions associated with topoisomerase III enzyme in other bacteria.
authors.affiliation: Structural Biology Center, Biosciences Division, Argonne National Laboratory, Argonne, IL 60439, USA. Electronic address: ktan@anl.gov.
authors.forename: Kemin
authors.initials: K
authors.lastname: Tan
authors.affiliation: Department of Chemistry and Biochemistry, Florida International University, Miami, FL 33199, USA.
authors.forename: Nan
authors.initials: N
authors.lastname: Cao
authors.affiliation: Department of Biochemistry and Molecular Biology, New York Medical College, Valhalla, NY 10595, USA.
authors.forename: Bokun
authors.initials: B
authors.lastname: Cheng
authors.affiliation: Structural Biology Center, Biosciences Division, Argonne National Laboratory, Argonne, IL 60439, USA.
authors.forename: Andrzej
authors.initials: A
authors.lastname: Joachimiak
authors.affiliation: Department of Chemistry and Biochemistry, Florida International University, Miami, FL 33199, USA; Biomolecular Sciences Institute, Florida International University, Miami, FL 33199, USA. Electronic address: ytsedinh@fiu.edu.
authors.forename: Yuk-Ching
authors.initials: YC
authors.lastname: Tse-Dinh
mesh_terms: D000595:Amino Acid Sequence; D018360:Crystallography, X-Ray; D004247:DNA / Q000378:metabolism*; D004264:DNA Topoisomerases, Type I / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000235:genetics; D011487:Protein Conformation; D016415:Sequence Alignment
chemical_list: D004247:DNA; D004264:DNA Topoisomerases, Type I
keywords: TB; TopA; crystallization; protein domains; protein fold