Ankyrin domains across the Tree of Life

Jernigan, Kristin K.; Bordenstein, Seth R.

doi:10.7717/peerj.264

Cited by 82 publications

(89 citation statements)

References 43 publications

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“…4,14,18,19 Little is known, however, regarding the physiological roles of ankyrin repeats in prokaryotes. Their prevalence in proteins from intracellular pathogens such as Legionella pneumophila is especially intriguing, however.…”

Section: Resultsmentioning

confidence: 99%

New Role for the Ankyrin Repeat Revealed by a Study of the N-Formyltransferase from Providencia alcalifaciens

2015

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N -formylated sugars such as 3,6-dideoxy-3-formamido-D-glucose (Qui3NFo) have been observed on the lipopolysaccharides of various pathogenic bacteria including Providencia alcalifaciens, a known cause of gastroenteritis. These unusual carbohydrates are synthesized in vivo as dTDP-linked sugars. The biosynthetic pathway for the production of dTDP-Qui3NFo requires five enzymes with the last step catalyzed by an N-formyltransferase that utilizes N10-tetrahydrofolate as a cofactor. Here we describe a structural and functional investigation of the P. alcalifaciens N-formyltransferase, hereafter referred to as QdtF. For this analysis the structure of the dimeric enzyme was solved in the presence of N5-formyltetrahydrofolate, a stable cofactor, and dTDP-3,6-dideoxy-3-amino-D-glucose (dTDP-Qui3N) to 1.5-Å resolution. The overall fold of the subunit consists of three regions with the N-terminal and middle motifs followed by an ankyrin repeat domain. Whereas the ankyrin repeat is a common eukaryotic motif involved in protein:protein interactions, reports of its presence in prokaryotic enzymes have been limited. Unexpectedly, this ankyrin repeat houses a second binding pocket for dTDP-Qui3N, which is characterized by extensive interactions between the protein and the ligand. To address the effects of this second binding site on catalysis, a site-directed mutant protein, W305A, was constructed. Kinetic analyses demonstrated that the catalytic activity of the W305A variant was reduced by approximately sevenfold. The structure of the W305A mutant protein in complex with N5-formyltetrahydrofolate and dTDP-Qui3N was subsequently determined to 1.5-Å resolution. The electron density map clearly showed that ligand binding had been completely abolished in the auxiliary pocket. The wild-type enzyme was also tested for activity against dTDP-3,6-dideoxy-3-amino-D-galactose (dTDP-Fuc3N) as a substrate. Strikingly, sigmoidal kinetics were observed indicating homotropic allosteric behavior. Although the identity of the ligand that regulates QdtF activity in vivo is presently unknown, our results still provide the first example of an ankyrin repeat functioning in small molecule binding.

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Section: Resultsmentioning

confidence: 99%

New Role for the Ankyrin Repeat Revealed by a Study of the N-Formyltransferase from Providencia alcalifaciens

2015

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“…The ankyrin repeat is one of the most common protein-protein interaction motifs in nature (26). Ankyrin repeat-containing proteins (Anks) are emerging as key virulence factors of intracellular bacteria and viruses for interacting with host cell proteins to coopt cellular processes (27).…”

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confidence: 99%

“…Ankyrin repeat-containing proteins (Anks) are emerging as key virulence factors of intracellular bacteria and viruses for interacting with host cell proteins to coopt cellular processes (27). O. tsutsugamushi encodes one of the largest known Ank repertoires of any microbe (26). Most O. tsutsugamushi Anks also carry an F-box domain that is capable of interacting with SKP1 of the SCF1 ubiquitin ligase complex (28,29), which normally functions in eukaryotic cells to tag proteins for degradation by the 26S proteasome (30).…”

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confidence: 99%

Orientia tsutsugamushi Modulates Endoplasmic Reticulum-Associated Degradation To Benefit Its Growth

et al. 2018

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Orientia tsutsugamushi, an obligate intracellular bacterium that is auxotrophic for the aromatic amino acids and histidine, causes scrub typhus, a potentially deadly infection that threatens 1 billion people. O. tsutsugamushi growth is minimal during the first 24 to 48 h of infection but its growth becomes logarithmic thereafter. How the pathogen modulates cellular functions to support its growth is poorly understood. The unfolded protein response (UPR) is a cytoprotective pathway that relieves endoplasmic reticulum (ER) stress by promoting ER-associated degradation (ERAD) of misfolded proteins. Here, we show that O. tsutsugamushi invokes the UPR in the first 48 h and benefits from ER stress in an amino acid-dependent manner. O. tsutsugamushi also impedes ERAD during this time period. By 72 h, ER stress is alleviated and ERAD proceeds unhindered. Sustained inhibition of ERAD using RNA interference results in an O. tsutsugamushi growth defect at 72 h that can be rescued by amino acid supplementation. Thus, O. tsutsugamushi temporally stalls ERAD until ERAD-derived amino acids are needed to support its growth. The O. tsutsugamushi effector Ank4 is linked to this phenomenon. Ank4 interacts with Bat3, a eukaryotic chaperone that is essential for ERAD, and is transiently expressed by O. tsutsugamushi during the infection period when it inhibits ERAD. Ectopically expressed Ank4 blocks ERAD to phenocopy O. tsutsugamushi infection. Our data reveal a novel mechanism by which an obligate intracellular bacterial pathogen modulates ERAD to satisfy its nutritional virulence requirements.

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“…Although few Anaplasmataceae effectors have been characterized, some encode ankyrin repeat motifs that are common in eukaryotes and may have implications for virulence 87 . Ankyrin repeats mediate protein–protein interactions, provide scaffolding for signalling pathways and localize molecules to subcellular compartments in eukaryotes 88 . Ankyrin-repeat containing proteins (Anks) in A. phagocytophilum and E. chaffeensis were shown to mediate host epigenetic changes to promote bacterial survival 69,89 .…”

Section: Discussionmentioning

confidence: 99%

Engineering of obligate intracellular bacteria: progress, challenges and paradigms

et al. 2017

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It is estimated that approximately one billion people are at risk of infection with obligate intracellular bacteria, but little is known about the underlying mechanisms that govern their life cycles. The difficulty in studying Chlamydia spp., Coxiella spp., Rickettsia spp., Anaplasma spp., Ehrlichia spp. and Orientia spp. is, in part, due to their genetic intractability Recently, genetic tools have been developed; however, optimizing the genomic manipulation of obligate intracellular bacteria remains challenging. In this Review, we describe the progress in, as well as the constraints that hinder, the systematic development of a genetic toolbox for obligate intracellular bacteria. We highlight how the use of genetically manipulated pathogens has facilitated a better understanding of microbial pathogenesis and immunity and how the engineering of obligate intracellular bacteria could enable the discovery of novel signalling circuits in host–pathogen interactions.

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Ankyrin domains across the Tree of Life

Cited by 82 publications

References 43 publications

New Role for the Ankyrin Repeat Revealed by a Study of the N-Formyltransferase from Providencia alcalifaciens

New Role for the Ankyrin Repeat Revealed by a Study of the N-Formyltransferase from Providencia alcalifaciens

Orientia tsutsugamushi Modulates Endoplasmic Reticulum-Associated Degradation To Benefit Its Growth

Engineering of obligate intracellular bacteria: progress, challenges and paradigms

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