The hexameric Rho motor is an RNA helicase that terminates transcription in bacteria

Rho is essential in many species

Rho is the target of the natural antibiotic bicyclomycin [BCM]

BCM prevents Rho ring closure around RNA and Rho motor activation

Structural basis for resistance of Mycobacterium tuberculosis Rho to bicyclomycin

Open (inactive) Closed (active)

E. coli Rho

James Berger lab

BCM

Structural basis for resistance of Mycobacterium tuberculosis Rho to bicyclomycin

Mtb Rho

Rho is essential in M. tuberculosis

MtbRho is resistant to bicyclomycin [BCM]

All residues contacting BCM in EcRho are strictly conserved in MtbRho

Skordalakes et al, & Berger, Structure (2005)

D’heygere et al., NAR (2015)

Structural basis for resistance of Mycobacterium tuberculosis Rho to bicyclomycin

Atomic model refined to 3.3 Å resolution

A Leu→Met mutation creates a steric bulk in the BCM binding pocket of MtbRho

Cryo-EM

Structural basis for resistance of Mycobacterium tuberculosis Rho to bicyclomycin

Reverting to a Leucine restores sensitivity to BCM

Also improves termination efficiency

Structural basis for resistance of Mycobacterium tuberculosis Rho to bicyclomycin

The M495L mutation stimulatesNTPase and helicase activities

Resistance to BCM by a bulkyLeu→Met substitution has a cost: reduced enzymatic efficiency

M495 R-loop

Structural basis for resistance of Mycobacterium tuberculosis Rho to bicyclomycin

ContributorsCBM, CNRS Orléans, France• Marc Boudvillain• Emmanuel Saridakis

(on sabattical leave from Greece)• Franck Coste• Isabelle Simon

CBS, CNRS Montpellier, France• Emmanuel Margeat• Rishi Vishwakarma• Josephine Lai-Kee-Him• Kevin Martin• Martin-Cohen-Gonsaud• Patrick Bron

The Leu→Met mutation conferring resistance to BCM is a taxa-specific trait

BCM-producing species (●) use a distinct mechanism of protection