Cambridge study shows possible path toward treating antibiotic-resistant tuberculosis

Tuberculosis is currently the deadliest infectious disease in the world, affecting nearly 2 billion people. It is caused by the organism Mycobacterium tuberculosis. 

The recent spread of antibiotic resistant strains of this bacteria has engendered research into new treatment methods. To reduce the risk of working with the deadly M. tuberculosis, much research has focused on the related, but non-pathogenic, species Mycobacterium smegmatis which has the additional advantage of fast reproduction time. 

A recent study by four University of Cambridge researchers, including Nobel prize winner John E. Walker, shows new features of the mechanism by which the bacterium synthesizes the crucial enzyme adenosine triphosphate (ATP). These results, give hope that new agents may be found to target the antibiotic-resistant strains of M. tuberculosis. 

Their study was published online Nov. 15, 2021, in the Proceedings of the National Academy of Sciences. 

ATP synthase inhibitors

In the United States, bedaquiline (Sirturo) has been used since 2013 to treat multidrug-resistant tuberculosis. The drug works by preventing the bacteria from synthesizing (ATP), the energy producing chemical needed by all life. ATP is generated in the bacterium by a molecular machine known as ATP synthase. 

But new strains resistant to bedaquiline have appeared. This new study elaborates features of the ATP synthase in Mycobacterium. smegmatis that are not present in human ATP synthase. The hope is that new drugs could be developed that could interrupt ATP synthesis in the bacteria without disrupting the same crucial process in humans.

New structures discovered

Using electron cryomicroscopy, the authors succeeded in describing three “highly significant attributes not recognized before that are crucial for understanding the mechanism and regulation of the mycobacterial enzyme," the researchers write. 

“First, we resolved not only the three main states in the catalytic cycle described before but also eight substates that portray structural and mechanistic changes occurring during a 360° catalytic cycle,” the researchers reported.

Second, they found that “a mechanism of auto-inhibition of ATP hydrolysis involves not only the engagement of the C-terminal region of an α-subunit in a loop in the γ-subunit, as proposed before, but also a ‘fail-safe’ mechanism involving the b′-subunit in the peripheral stalk that enhances engagement.”

"A third unreported characteristic,” the authors report, “is that the fused δ-subunit contains a duplicated domain in its N-terminal region where the two copies of the domain participate in similar modes of attachment of the two of three N-terminal regions of the α-subunits.”

(The regions they reference are illustrated in the graphic above.) 

All three of these newly defined features––the auto-inhibitory, the fail-safe mechanism, and the methods of attachment of the alpha subunits–– “provide targets for development of innovative anti-tubercular drugs,” the authors note.

The new work also confirms the binding site of the drug bedaquiline that had been described previously. 

Taken together these findings “have the potential to be exploited in the design of new inhibitors that could be developed into novel drugs against TB,” the authors note. 

New drug possibilities

The authors elaborated on possibilities for drug development. 

“The first of these sites involves the ‘hook’ and ‘catch’ features involved in the ATP hydrolytic inhibitory mechanism, described also in the prior structure," they wrote. “The second is the ‘fail-safe’ device that is likely to enhance the engagement of the hook and loop in state S1. The third is the unique mode of association of the PS with the N-terminal regions of the α-subunits. The ‘hook and catch’ device could be locked in place by small molecules that bind to both features, thereby preventing synthesis of ATP.”

The authors also offer a proposal.  

“The ‘fail-safe’ interaction could either be augmented by a small molecule binding across the feature, or its formation could be impeded by small molecules binding to its two elements," they wrote. "Compounds with the features of suitably designed ‘molecular glues’ might enhance the stability of both features and be effective inhibitors of ATP synthesis by the enzyme. The third target would require small molecules that would impede the attachment of the PS to the F1-domain during its assembly, thereby preventing the coupling of the pmf [proton motor force] to the synthesis of ATP. 

“Once such inhibitors had been identified, then would follow the extensive process of converting them into effective drugs for treating TB,” they concluded.

The authors of the study are Martin Montgomery, Jessica Petri, Tobias Spikes, and Nobel laureate Walker, all from the University of Cambridge in England. 

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M. Montgomery et al, Structure of the ATP synthase from Mycobacterium smegmatis provides targets for treating tuberculosis. PNAS (2021).

DOI: https://doi.org/10.1073/pnas.2111899118