Structural basis of protein arginine rhamnosylation by glycosyltransferase EarP

Sengoku, Toru; Suzuki, Takehiro; Dohmae, Naoshi; Watanabe, Chiduru; Honma, Teruki; Hikida, Yasushi; Yamaguchi, Yoshiki; Takahashi, Hideyuki; Yokoyama, Shigeyuki; Yanagisawa, Takatoshi

doi:10.1038/s41589-018-0002-y

Cited by 20 publications

(40 citation statements)

References 44 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The buried surface area between EarP and domain I of EF-P was 1,030 Å 2 provided by PISA. This is similar to the interface areas of 897.5 Å 2 between N. meningitidis EarP and EF-P (30) and 973 Å 2 between Escherichia coli EF-P and its modification enzyme GenX/EpmA (39).…”

Section: Resultssupporting

confidence: 76%

“…S4C). The rhamnose-accommodating cavity in our structure provides more hydrogen bonds than that observed for N. meningitidis EarP (30). In detail (Fig.…”

Section: Resultsmentioning

confidence: 74%

“…The ternary complex structure of P. aeruginosa EarP with TDP and EF-P shows that Asp12, Asp16, and Tyr112 interact with the acceptor Arg32* -nitrogen atoms directly. Asp16 of P. aeruginosa EarP corresponds to Asp20 of N. meningitidis EarP, which has been reported as the general base (30). Moreover, replacement of Glu273 of P. putida EarP, corresponding to Glu272 of P. aeruginosa EarP, with a glutamine resulted in undetectable EF-P rhamnosylation in vivo (29).…”

Section: Resultsmentioning

confidence: 92%

“…In contrast, in the structure of N. meningitidis EarP bound with TDP-Rha, the ␤-phosphate lies farther away from helix ␣1 (Fig. S4C), forming water-mediated hydrogen bonds with helix ␣1 (30).…”

Section: Resultsmentioning

confidence: 96%

“…Nevertheless, this structure missed structural elements in the N-terminal domain (NTD). T. Sengoku et al determined the crystal structures of EarP from N. meningitidis in the apo-and TDP-Rhabound forms, as well as in complex with domain I of EF-P (30). These researchers showed that EarP binds the entire ␤-sheet structure of EF-P domain I and recognizes its conserved residues through numerous side chain-specific interactions.…”

mentioning

confidence: 99%

See 4 more Smart Citations

Complex Structure of Pseudomonas aeruginosa Arginine Rhamnosyltransferase EarP with Its Acceptor Elongation Factor P

Liu

et al. 2019

J Bacteriol

View full text Add to dashboard Cite

A bacterial inverting glycosyltransferase EarP transfers rhamnose from dTDP-β-l-rhamnose (TDP-Rha) to Arg32 of translation elongation factor P (EF-P) to activate its function. We report here the structural and biochemical characterization of Pseudomonas aeruginosa EarP. In contrast to recently reported Neisseria meningitidis EarP, P. aeruginosa EarP exhibits differential conformational changes upon TDP-Rha and EF-P binding. Sugar donor binding enhances acceptor binding to EarP, as revealed by structural comparison between the apo-, TDP-Rha-, and TDP/EF-P-bound forms and isothermal titration calorimetry experiments. In vitro EF-P rhamnosylation combined with active-site geometry indicates that Asp16 corresponding to Asp20 of N. meningitidis EarP is the catalytic base, whereas Glu272 is another putative catalytic residue. Our study should provide the basis for EarP-targeted inhibitor design against infections from P. aeruginosa and other clinically relevant species. IMPORTANCE Posttranslational rhamnosylation of EF-P plays a key role in Pseudomonas aeruginosa, establishing virulence and antibiotic resistance, as well as survival. The detailed structural and biochemical characterization of the EF-P-specific rhamnosyltransferase EarP from P. aeruginosa not only demonstrates that sugar donor TDP-Rha binding enhances acceptor EF-P binding to EarP but also should provide valuable information for the structure-guided development of its inhibitors against infections from P. aeruginosa and other EarP-containing pathogens.

show abstract

Section: Resultssupporting

confidence: 76%

“…S4C). The rhamnose-accommodating cavity in our structure provides more hydrogen bonds than that observed for N. meningitidis EarP (30). In detail (Fig.…”

Section: Resultsmentioning

confidence: 74%

Section: Resultsmentioning

confidence: 92%

Section: Resultsmentioning

confidence: 96%

mentioning

confidence: 99%

See 3 more Smart Citations

Complex Structure of Pseudomonas aeruginosa Arginine Rhamnosyltransferase EarP with Its Acceptor Elongation Factor P

Liu

et al. 2019

J Bacteriol

View full text Add to dashboard Cite

show abstract

Biochemical characterization of a glycosyltransferase Gtf3 from Mycobacterium smegmatis: a case study of improved protein solubilization

et al. 2020

View full text Add to dashboard Cite

Glycosyltransferases (GTs) are widely present in several organisms. These enzymes specifically transfer sugar moieties to a range of substrates. The processes of bacterial glycosylation of the cell wall and their relations with host-pathogen interactions have been studied extensively, yet the majority of mycobacterial GTs involved in the cell wall synthesis remain poorly characterized. Glycopeptidolipids (GPLs) are major class of glycolipids present on the cell wall of various mycobacterial species. They play an important role in drug resistance and host-pathogen interaction virulence. Gtf3 enzyme performs a key step in the biosynthesis of triglycosylated GPLs. Here, we describe a general procedure to achieve expression, purification, and crystallization of recombinant protein Gtf3 from Mycobacterium smegmatis using an E. coli expression system. We reported also a combined bioinformatics and biochemical methods to predict aggregation propensity and improve protein solubilization of recombinant Gtf3. NVoy, a carbohydrate-based polymer reagent, was added to prevent protein aggregation by binding to hydrophobic protein surfaces of Gtf3. Using intrinsic tryptophan fluorescence quenching experiments, we also demonstrated that Gtf3-NVoy enzyme interacted with TDP and UDP nucleotide ligands. This case report proposes useful tools for the study of other glycosyltransferases which are rather difficult to characterize and crystallize.

show abstract

Translation factor accelerating peptide bond formation on the ribosome: EF-P and eIF5A as entropic catalysts and a potential drug targets

Mudryi¹,

Peske²,

Rodnina³

2023

BBA Advances

View full text Add to dashboard Cite

Structural basis of protein arginine rhamnosylation by glycosyltransferase EarP

Cited by 20 publications

References 44 publications

Complex Structure of Pseudomonas aeruginosa Arginine Rhamnosyltransferase EarP with Its Acceptor Elongation Factor P

Complex Structure of Pseudomonas aeruginosa Arginine Rhamnosyltransferase EarP with Its Acceptor Elongation Factor P

Biochemical characterization of a glycosyltransferase Gtf3 from Mycobacterium smegmatis: a case study of improved protein solubilization

Translation factor accelerating peptide bond formation on the ribosome: EF-P and eIF5A as entropic catalysts and a potential drug targets

Contact Info

Product

Resources

About