Overexpression, purification, enzymatic and microscopic characterization of recombinant mycobacterial F-ATP synthase

Overexpression, purification, enzymatic and microscopic characterization of recombinant mycobacterial F-ATP synthase

The F-ATP synthase is an essential enzyme in mycobacteria, including the pathogenic Mycobacterium tuberculosis. Several new compounds in the TB-drug pipeline target the F-ATP synthase. In light of the importance and pharmacological attractiveness of this novel antibiotic target, tools have to be dev...

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Bibliographic Details
Main Authors: Saw, Wuan-Geok, Wong, Chui-Fann, Dick, Thomas, Grüber, Gerhard
Other Authors: School of Biological Sciences
Format: Article
Language:English
Published: 2020
Subjects:
Science::Biological sciences::Biochemistry
Science::Biological sciences::Molecular biology
F-ATP Synthase
Bioenergetics
Online Access:https://hdl.handle.net/10356/139317
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Institution: Nanyang Technological University
Language: English
Description
Summary:The F-ATP synthase is an essential enzyme in mycobacteria, including the pathogenic Mycobacterium tuberculosis. Several new compounds in the TB-drug pipeline target the F-ATP synthase. In light of the importance and pharmacological attractiveness of this novel antibiotic target, tools have to be developed to generate a recombinant mycobacterial F1FO ATP synthase to achieve atomic insight and mutants for mechanistic and regulatory understanding as well as structure-based drug design. Here, we report the first genetically engineered, purified and enzymatically active recombinant M. smegmatis F1FO ATP synthase. The projected 2D- and 3D structures of the recombinant enzyme derived from negatively stained electron micrographs are presented. Furthermore, the first 2D projections from cryo-electron images are revealed, paving the way for an atomic resolution structure determination.

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