Structures and transport dynamics of a Campylobacter jejuni multidrug efflux pump

Su, Chih‐Chia; Yin, Linxiang; Kumar, Nitin; Dai, Lei; Radhakrishnan, Abhijith; Bolla, Jani Reddy; Lei, Hanlun; Chou, Tsung‐Han; Delmar, Jared A.; Rajashankar, Kanagalaghatta R.; Zhang, Qijing; Shin, Yeon‐Kyun; Yu, Edward

doi:10.1038/s41467-017-00217-z

Cited by 76 publications

(150 citation statements)

References 43 publications

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“…S3). Although MmpL3 belongs to a subclass of the RND superfamily, its 3D topology is unique and very different from the existing structures of RND transporters, including AcrB (24,25), MexB (26), CusA (27,28), MtrD (29), CmeB (30), and HpnN (31). Thus, the available structural information of RND proteins cannot be used to understand the function of this transporter.…”

Section: Resultsmentioning

confidence: 99%

“…Its oligomerization state and 3D structure are very distinct from all other known structures belonging to the RND superfamily of membrane proteins. For example, the well-studied hydrophobic and amphiphilic efflux subfamily of RND transporters, including AcrB (24,25), MexB (26), MtrD (29), and CmeB (30), exist as trimers within the cytoplasmic membrane of Gram-negative bacteria. They have large, periplasmic domains that confer substrate specificity and interact with periplasmic adaptors and outer membrane channels to form tripartite efflux complexes.…”

Section: Discussionmentioning

confidence: 99%

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MmpL3 is a lipid transporter that binds trehalose monomycolate and phosphatidylethanolamine

Klenotic

Bolla

et al. 2019

Proc. Natl. Acad. Sci. U.S.A.

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115

146

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The cell envelope of Mycobacterium tuberculosis is notable for the abundance of mycolic acids (MAs), essential to mycobacterial viability, and of other species-specific lipids. The mycobacterial cell envelope is extremely hydrophobic, which contributes to virulence and antibiotic resistance. However, exactly how fatty acids and lipidic elements are transported across the cell envelope for cellwall biosynthesis is unclear. Mycobacterial membrane protein Large 3 (MmpL3) is essential and required for transport of trehalose monomycolates (TMMs), precursors of MA-containing trehalose dimycolates (TDM) and mycolyl arabinogalactan peptidoglycan, but the exact function of MmpL3 remains elusive. Here, we report a crystal structure of Mycobacterium smegmatis MmpL3 at a resolution of 2.59 Å, revealing a monomeric molecule that is structurally distinct from all known bacterial membrane proteins. A previously unknown MmpL3 ligand, phosphatidylethanolamine (PE), was discovered inside this transporter. We also show, via native mass spectrometry, that MmpL3 specifically binds both TMM and PE, but not TDM, in the micromolar range. These observations provide insight into the function of MmpL3 and suggest a possible role for this protein in shuttling a variety of lipids to strengthen the mycobacterial cell wall. mycobacterial membrane protein Large | cell-wall biogenesis | MmpL3 transporter | X-ray crystallography | native mass spectrometry

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

MmpL3 is a lipid transporter that binds trehalose monomycolate and phosphatidylethanolamine

Klenotic

Bolla

et al. 2019

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

115

146

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“…177,178 Interestingly, for CmeB (an AcrB homolog from C. jejuni), it was proposed that individual protomers operate independently of each other. 179 However, the examination of the isolated IMP component might not necessarily resemble the conformational dynamics within the assembled tripartite complex.…”

Section: Remote Alternative Access Drug/proton Antiport In Acrbmentioning

confidence: 99%

AcrB: a mean, keen, drug efflux machine

Kobylka

Kuth

Müller

et al. 2019

Annals of the New York Academy of Sciences

123

135

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Gram-negative bacteria are intrinsically resistant against cytotoxic substances by means of their outer membrane and a network of multidrug efflux systems, acting in synergy. Efflux pumps from various superfamilies with broad substrate preferences sequester and pump drugs across the inner membrane to supply the highly polyspecific and powerful tripartite resistance-nodulation-cell division (RND) efflux pumps with compounds to be extruded across the outer membrane barrier. In Escherichia coli, the tripartite efflux system AcrAB-TolC is the archetype RND multiple drug efflux pump complex. The homotrimeric inner membrane component acriflavine resistance B (AcrB) is the drug specificity and energy transduction center for the drug/proton antiport process. Drugs are bound and expelled via a cycle of mainly three consecutive states in every protomer, constituting a flexible alternating access channel system. This review recapitulates the molecular basis of drug and inhibitor binding, including mechanistic insights into drug efflux by AcrB. It also summarizes 17 years of mutational analysis of the gene acrB, reporting the effect of every substitution on the ability of E. coli to confer resistance toward antibiotics (http://goethe.link/ AcrBsubstitutions). We emphasize the functional robustness of AcrB toward single-site substitutions and highlight regions that are more sensitive to perturbation.

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“…CusA (27,28), MtrD (29), CmeB (30) and HpnN (31). Thus, the available structural information of RND proteins cannot be used to understand the function of this transporter.…”

Section: Overall Structure Of the M Smegmatis Mmpl3 Transportermentioning

confidence: 99%

MmpL3 is a lipid transporter that binds trehalose monomycolate and phosphatidylethanolamine

Klenotic

Bolla

et al. 2019

Preprint

Self Cite

View full text Add to dashboard Cite

The cell envelope of Mycobacterium tuberculosis is notable for the abundance of mycolic acids (MAs), which are essential to mycobacterial viability, and other species-specific lipids. The mycobacterial cell envelope is extremely hydrophobic, contributes to virulence and antibiotic resistance. Yet, exactly how fatty acids and lipidic elements are transported across the cell envelope for cell wall biosynthesis is unclear. Mycobacterial membrane protein Large 3 (MmpL3) is essential and required for transport of trehalose monomycolates (TMMs), precursors of MA containing trehalose dimycolates (TDM) and mycolyl arabinogalactan peptidoglycan (mAGP), but the exact function of MmpL3 remains elusive. Here, we report a high-resolution crystal structure of M. smegmatis MmpL3, revealing a monomeric molecule that is structurally distinct from all known bacterial membrane proteins. A previously unknown MmpL3 ligand, phosphatidylethanolamine (PE), was discovered inside this transporter. We also show, via native mass spectrometry, that MmpL3 specifically binds both TMM and PE, but not TDM, in the micromolar range. These observations provide insight into the function of MmpL3 and suggest a possible role for this protein in shuttling a variety of lipids to strengthen the mycobacterial cell wall.

show abstract

Structures and transport dynamics of a Campylobacter jejuni multidrug efflux pump

Cited by 76 publications

References 43 publications

MmpL3 is a lipid transporter that binds trehalose monomycolate and phosphatidylethanolamine

MmpL3 is a lipid transporter that binds trehalose monomycolate and phosphatidylethanolamine

AcrB: a mean, keen, drug efflux machine

MmpL3 is a lipid transporter that binds trehalose monomycolate and phosphatidylethanolamine

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