Molecular mechanism underlying promiscuous polyamine recognition by spermidine acetyltransferase

Sugiyama, Shigeru; Ishikawa, Sae; Tomitori, Hideyuki; Niiyama, Mayumi; Hirose, Mika; Miyazaki, Yuma; Higashi, Kyohei; Murata, Michio; Adachi, Hiroaki; Takano, Kazufumi; Murakami, Satoshi; Inoue, Tsuyoshi; Mori, Yusuke; Kashiwagi, Keiko; Igarashi, Kazuei; Matsumura, Hideki

doi:10.1016/j.biocel.2016.05.003

Cited by 10 publications

(24 citation statements)

References 36 publications

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“…Therefore, residues within the WFEE portion of the allosteric loop are critical for activity, but we do not know whether all residues or only a subset of these four residues are required for activity toward spm for the VcSpeG enzyme. These four residues are conserved across all structurally characterized SpeG enzymes, and Sugiyama et al previously showed point mutants of the E. coli SpeG enzyme in this comparable region were critical for activity toward spd (Sugiyama et al, 2016). However, we currently do not know whether the VcSpeG allosteric site must bind polyamine for turnover to occur and which residues in the allosteric site might transmit this signal when spm or spd is bound.…”

Section: Two Previously Unexplored Regions Of Vcspeg Are Critical For Activitymentioning

confidence: 93%

The Vibrio cholerae SpeG Spermidine/Spermine N-Acetyltransferase Allosteric Loop and β6-β7 Structural Elements Are Critical for Kinetic Activity

Tsimbalyuk

Lim

et al. 2021

Front. Mol. Biosci.

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Polyamines regulate many important biological processes including gene expression, intracellular signaling, and biofilm formation. Their intracellular concentrations are tightly regulated by polyamine transport systems and biosynthetic and catabolic pathways. Spermidine/spermine N-acetyltransferases (SSATs) are catabolic enzymes that acetylate polyamines and are critical for maintaining intracellular polyamine homeostasis. These enzymes belong to the Gcn5-related N-acetyltransferase (GNAT) superfamily and adopt a highly conserved fold found across all kingdoms of life. SpeG is an SSAT protein found in a variety of bacteria, including the human pathogen Vibrio cholerae. This protein adopts a dodecameric structure and contains an allosteric site, making it unique compared to other SSATs. Currently, we have a limited understanding of the critical structural components of this protein that are required for its allosteric behavior. Therefore, we explored the importance of two key regions of the SpeG protein on its kinetic activity. To achieve this, we created various constructs of the V. cholerae SpeG protein, including point mutations, a deletion, and chimeras with residues from the structurally distinct and non-allosteric human SSAT protein. We measured enzyme kinetic activity toward spermine for ten constructs and crystallized six of them. Ultimately, we identified specific portions of the allosteric loop and the β6-β7 structural elements that were critical for enzyme kinetic activity. These results provide a framework for further study of the structure/function relationship of SpeG enzymes from other organisms and clues toward the structural evolution of members of the GNAT family across domains of life.

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Section: Two Previously Unexplored Regions Of Vcspeg Are Critical For Activitymentioning

confidence: 93%

The Vibrio cholerae SpeG Spermidine/Spermine N-Acetyltransferase Allosteric Loop and β6-β7 Structural Elements Are Critical for Kinetic Activity

Tsimbalyuk

Lim

et al. 2021

Front. Mol. Biosci.

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“…Studies indicate that SpeG plays an important role in virulence and pathogenicity of many bacteria, which have evolved SpeG-dependent strategies to control polyamine concentrations and survive in their hosts [3-6]. Kinetic and structural analyses have demonstrated that SpeG from both Escherichia coli and Vibrio cholerae can acetylate spermidine [7-9]. These studies also showed that SpeG from V. cholerae is an allosteric protein; when spermidine binds to its allosteric site, SpeG exhibits a symmetric closed dodecameric structure [7, 9].…”

Section: Introductionmentioning

confidence: 99%

“…Kinetic and structural analyses have demonstrated that SpeG from both Escherichia coli and Vibrio cholerae can acetylate spermidine [7-9]. These studies also showed that SpeG from V. cholerae is an allosteric protein; when spermidine binds to its allosteric site, SpeG exhibits a symmetric closed dodecameric structure [7, 9]. Finally, in the absence of spermidine binding, V. cholerae SpeG can adopt a unique asymmetric dodecameric structure with an open conformational state [10].…”

Section: Introductionmentioning

confidence: 99%

The spermidine acetyltransferase SpeG regulates transcription of the small RNA RprA

Hu¹,

Filippova

Dang

et al. 2018

Preprint

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28Spermidine N-acetyltransferase (SpeG) acetylates and thus neutralizes toxic 29 polyamines. Studies indicate that SpeG plays an important role in virulence and 30 pathogenicity of many bacteria, which have evolved SpeG-dependent strategies 31 to control polyamine concentrations and survive in their hosts. In Escherichia coli, 32 the two-component response regulator RcsB is reported to be subject to N ε -33 acetylation on several lysine residues, resulting in reduced DNA binding affinity 34 and reduced transcription of the small RNA rprA; however, the physiological 35 acetylation mechanism responsible for this behavior has not been fully determined. 36Here, we performed an acetyltransferase screen and found that SpeG inhibits rprA 37 promoter activity in an acetylation-independent manner. Surface plasmon 38 resonance analysis revealed that SpeG can physically interact with the DNA-39 binding carboxyl domain of RcsB. We hypothesize that SpeG interacts with the 40 DNA-binding domain of RcsB and that this interaction might be responsible for 41SpeG-dependent inhibition of RcsB-dependent rprA transcription. This work 42 provides a model for SpeG as a modulator of E. coli transcription through its ability 43 to interact with the transcription factor RcsB. This is the first study to provide 44 evidence that an enzyme involved in polyamine metabolism can influence the 45 function of the global regulator RcsB, which integrates information concerning 46 envelope stresses and central metabolic status to regulate diverse behaviors. 47

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“…Studies indicate that SpeG plays an important role in virulence and pathogenicity of many bacteria, which have evolved SpeG-dependent strategies to control polyamine concentrations and survive in their hosts [3–6]. Kinetic and structural analyses have demonstrated that SpeG from both Escherichia coli and Vibrio cholerae can acetylate spermidine [7–9]. These studies also showed that SpeG from V .…”

Section: Introductionmentioning

confidence: 99%

“…These studies also showed that SpeG from V . cholerae is an allosteric protein; when spermidine binds to its allosteric site, SpeG exhibits a symmetric closed dodecameric structure [7, 9]. Finally, in the absence of spermidine binding, V .…”

Section: Introductionmentioning

confidence: 99%

The spermidine acetyltransferase SpeG regulates transcription of the small RNA rprA

Filippova

Dang

et al. 2018

PLoS ONE

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Spermidine N-acetyltransferase (SpeG) acetylates and thus neutralizes toxic polyamines. Studies indicate that SpeG plays an important role in virulence and pathogenicity of many bacteria, which have evolved SpeG-dependent strategies to control polyamine concentrations and survive in their hosts. In Escherichia coli, the two-component response regulator RcsB is reported to be subject to Nε-acetylation on several lysine residues, resulting in reduced DNA binding affinity and reduced transcription of the small RNA rprA; however, the physiological acetylation mechanism responsible for this behavior has not been fully determined. Here, we performed an acetyltransferase screen and found that SpeG inhibits rprA promoter activity in an acetylation-independent manner. Surface plasmon resonance analysis revealed that SpeG can physically interact with the DNA-binding carboxyl domain of RcsB. We hypothesize that SpeG interacts with the DNA-binding domain of RcsB and that this interaction might be responsible for SpeG-dependent inhibition of RcsB-dependent rprA transcription. This work provides a model for SpeG as a modulator of E. coli transcription through its ability to interact with the transcription factor RcsB. This is the first study to provide evidence that an enzyme involved in polyamine metabolism can influence the function of the global regulator RcsB, which integrates information concerning envelope stresses and central metabolic status to regulate diverse behaviors.

show abstract

Molecular mechanism underlying promiscuous polyamine recognition by spermidine acetyltransferase

Cited by 10 publications

References 36 publications

The Vibrio cholerae SpeG Spermidine/Spermine N-Acetyltransferase Allosteric Loop and β6-β7 Structural Elements Are Critical for Kinetic Activity

The Vibrio cholerae SpeG Spermidine/Spermine N-Acetyltransferase Allosteric Loop and β6-β7 Structural Elements Are Critical for Kinetic Activity

The spermidine acetyltransferase SpeG regulates transcription of the small RNA RprA

The spermidine acetyltransferase SpeG regulates transcription of the small RNA rprA

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