Characterizing septum inhibition in Mycobacterium tuberculosis for novel drug discovery.
A temperature sensitive mutation in the cell division protein FtsZ was used in combination with transcriptional analysis to identify biomarkers for inhibition of septum formation. Crystallography and modeling revealed that the glycine for aspartate substitution at amino acid 210 was located in helix 8 of the protein, adjacent to the T7 synergy loop. To verify the molecular behavior of FtsZ D210G, the in vitro activity and structural stability were evaluated as a function of temperature. These analyses confirmed that the FtsZ D210G mutant had reduced GTPase and polymerization activity compared to wild-type FtsZ, and CD spectroscopy demonstrated that both FtsZ D210G and wild-type FtsZ had similar structure and stability. Significantly, the FtsZ D210G merodiploid strain of M. tuberculosis had compromised growth at 37 degrees C, substantiating the suitability of FtsZ D210G as a molecular tool for global analysis in response to improper FtsZ polymerization and septum inhibition. Advanced model-based bioinformatics and transcriptional mapping were used to identify high-content multiple features that provide biomarkers for the development of a rational drug screening platform for discovering novel chemotherapeutics that target cell division.
Respicio
Laurel
L
Rocky Mountain Regional Center of Excellence, Mycobacteria Research Laboratories, Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO 80523, United States.
Nair
Pravin A
PA
Huang
Qing
Q
Anil
Burcu
B
Tracz
Sylvia
S
Truglio
James J
JJ
Kisker
Caroline
C
Raleigh
Daniel P
DP
Ojima
Iwao
I
Knudson
Dennis L
DL
Tonge
Peter J
PJ
Slayden
Richard A
RA
D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000737:chemistry; D002453:Cell Cycle; D002455:Cell Division; D002942:Circular Dichroism / Q000379:methods; D018360:Crystallography, X-Ray / Q000379:methods; D003598:Cytoskeletal Proteins / Q000096:biosynthesis* / Q000737:chemistry; D020558:GTP Phosphohydrolases / Q000096:biosynthesis*; D015964:Gene Expression Regulation, Bacterial; D006801:Humans; D016297:Mutagenesis, Site-Directed; D009169:Mycobacterium tuberculosis / Q000235:genetics* / Q000502:physiology; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D003598:Cytoskeletal Proteins; C071525:FtsZ protein, Bacteria; D020558:GTP Phosphohydrolases