Small-Molecule Acetylation by GCN5-Related
            <i>N</i>
            -Acetyltransferases in Bacteria

Burckhardt, Rachel M.; Escalante‐Semerena, Jorge C.

doi:10.1128/mmbr.00090-19

Cited by 33 publications

(35 citation statements)

References 253 publications

(444 reference statements)

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“…Despite the structural and functional diversity of GNAT superfamily members, the catalytic and kinetic mechanisms of GNATs in the majority of literature reports are largely presumed to be well-established and nearly uniform. The canonical catalytic mechanism for GNATs is a general acid/base-catalyzed mechanism, while the kinetic mechanism is a direct acetyl-transfer mechanism ( Siehl et al, 2007 ; Vetting et al, 2008b ; Favrot et al, 2016 ; Burckhardt and Escalante-Semerena, 2020 ). Contrary to this perspective, several members of the GNAT superfamily actually utilize a range of catalytic mechanisms.…”

Section: Discussionmentioning

confidence: 99%

“…Acetyltransferases are fascinating enzymes found across all domains of life. They are critically important for various cellular functions including those of anabolic and catabolic pathways, cell wall modification, xenobiotic metabolism, and antibiotic drug resistance ( Sim et al, 2014 ; Hentchel and Escalante-Semerena, 2015 ; Zhang et al, 2017 ; Burckhardt and Escalante-Semerena, 2020 ; Sharma et al, 2020 ). Their seemingly simple ability to catalyze the transfer of an acetyl moiety from a donor molecule to an acceptor molecule is compounded by the diversity of structural scaffolds, active site residues, and kinetic mechanisms that they utilize.…”

Section: Introductionmentioning

confidence: 99%

“…While the donor site is relatively conserved, the residues that comprise the acceptor site vary widely and contribute to substrate specificity. The majority of GNATs that have been functionally characterized perform N- acetylation of primary amines ( Vetting et al, 2005b ; Favrot et al, 2016 ; Burckhardt and Escalante-Semerena, 2020 ), but a few examples of O- acetylation of hydroxyl groups exist ( Daigle et al, 1999 ; Hegde et al, 2001 ). Recent studies have also shown that the GNAT fold has been repurposed by some organisms to catalyze decarboxylation instead of acyl transfer, thus highlighting the sheer diversity of reaction capabilities of members of this superfamily of proteins ( Skiba et al, 2020 ).…”

Section: Introductionmentioning

confidence: 99%

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Gcn5-Related N-Acetyltransferases (GNATs) With a Catalytic Serine Residue Can Play Ping-Pong Too

Baumgartner

Mohammad

Czub

et al. 2021

Front. Mol. Biosci.

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Enzymes in the Gcn5-related N-acetyltransferase (GNAT) superfamily are widespread and critically involved in multiple cellular processes ranging from antibiotic resistance to histone modification. While acetyl transfer is the most widely catalyzed reaction, recent studies have revealed that these enzymes are also capable of performing succinylation, condensation, decarboxylation, and methylcarbamoylation reactions. The canonical chemical mechanism attributed to GNATs is a general acid/base mechanism; however, mounting evidence has cast doubt on the applicability of this mechanism to all GNATs. This study shows that the Pseudomonas aeruginosa PA3944 enzyme uses a nucleophilic serine residue and a hybrid ping-pong mechanism for catalysis instead of a general acid/base mechanism. To simplify this enzyme’s kinetic characterization, we synthesized a polymyxin B substrate analog and performed molecular docking experiments. We performed site-directed mutagenesis of key active site residues (S148 and E102) and determined the structure of the E102A mutant. We found that the serine residue is essential for catalysis toward the synthetic substrate analog and polymyxin B, but the glutamate residue is more likely important for substrate recognition or stabilization. Our results challenge the current paradigm of GNAT mechanisms and show that this common enzyme scaffold utilizes different active site residues to accomplish a diversity of catalytic reactions.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Gcn5-Related N-Acetyltransferases (GNATs) With a Catalytic Serine Residue Can Play Ping-Pong Too

Baumgartner

Mohammad

Czub

et al. 2021

Front. Mol. Biosci.

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“…How then would the AT domain act to inhibit reversals? Acetyltransferases of the GNAT family can acetylate amino groups of different substrates including small molecules, peptidoglycan, nucleotide-linked monosaccharides and proteins; however, the substrate cannot be determined based on protein sequence (80, 81). Therefore, further studies are required to determine the substrate of PixB.…”

Section: Discussionmentioning

confidence: 99%

Three PilZ domain proteins, PlpA, PixA and PixB, have distinct functions in regulation of motility and development inMyxococcus xanthus

Kuzmich

Skotnicka

Szadkowski

et al. 2021

Preprint

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In bacteria, the nucleotide-based second messenger bis-(3'-5')-cyclic dimeric GMP (c-di-GMP) binds to effectors to generate outputs in response to changes in the environment. In Myxococcus xanthus, c-di-GMP regulates type IV pili-dependent motility and the starvation-induced developmental program that results in the formation of spore-filled fruiting bodies; however, little is known about the effectors that bind c-di-GMP. Here, we systematically inactivated all 24 genes encoding PilZ domain-containing proteins, which are among the most common c-di-GMP receptors. We confirm that PlpA, a stand-alone PilZ-domain protein, is specifically important for motility and that Pkn1, which is composed of a Ser/Thr domain and a PilZ domain, is specifically important for development. Moreover, we identify two PilZ-domain proteins that have distinct functions in regulating motility and development. PixB, which is composed of two PilZ domains and an acetyltransferase domain, binds c-di-GMP in vitro and regulates type IV pili-dependent and gliding motility upstream of the Frz chemosensory system as well as development. The acetyltransferase domain is required and sufficient for function during growth while all three domains and c-di-GMP binding are essential for PixB function during development. PixA is a response regulator composed of a PilZ domain and a receiver domain, binds c-di-GMP in vitro, and regulates motility downstream of the Frz chemosensory system by setting up the polarity of the two motility systems. Our results support a model whereby the three proteins PlpA, PixA and PixB act in parallel pathways and have distinct functions to regulation of motility.

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“…This GNAT appears to be the first enzymatic acylation mechanism with ability to modify polymyxin. Consequently, some recent work has been carried out on GNAT inhibitors [ 33 ]. However, it is not known if such enzymatic modification may confer resistance to colistin.…”

Section: From Electron Microscopy To the Discovery Of Regulatory Gmentioning

confidence: 99%

The History of Colistin Resistance Mechanisms in Bacteria: Progress and Challenges

2021

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Since 2015, the discovery of colistin resistance genes has been limited to the characterization of new mobile colistin resistance (mcr) gene variants. However, given the complexity of the mechanisms involved, there are many colistin-resistant bacterial strains whose mechanism remains unknown and whose exploitation requires complementary technologies. In this review, through the history of colistin, we underline the methods used over the last decades, both old and recent, to facilitate the discovery of the main colistin resistance mechanisms and how new technological approaches may help to improve the rapid and efficient exploration of new target genes. To accomplish this, a systematic search was carried out via PubMed and Google Scholar on published data concerning polymyxin resistance from 1950 to 2020 using terms most related to colistin. This review first explores the history of the discovery of the mechanisms of action and resistance to colistin, based on the technologies deployed. Then we focus on the most advanced technologies used, such as MALDI-TOF-MS, high throughput sequencing or the genetic toolbox. Finally, we outline promising new approaches, such as omics tools and CRISPR-Cas9, as well as the challenges they face. Much has been achieved since the discovery of polymyxins, through several innovative technologies. Nevertheless, colistin resistance mechanisms remains very complex.

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Small-Molecule Acetylation by GCN5-Related N -Acetyltransferases in Bacteria

Cited by 33 publications

References 253 publications

Gcn5-Related N-Acetyltransferases (GNATs) With a Catalytic Serine Residue Can Play Ping-Pong Too

Gcn5-Related N-Acetyltransferases (GNATs) With a Catalytic Serine Residue Can Play Ping-Pong Too

Three PilZ domain proteins, PlpA, PixA and PixB, have distinct functions in regulation of motility and development inMyxococcus xanthus

The History of Colistin Resistance Mechanisms in Bacteria: Progress and Challenges

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