Gcn5-Related N-Acetyltransferases (GNATs) With a Catalytic Serine Residue Can Play Ping-Pong Too
Copyright © 2021 Baumgartner, Habeeb Mohammad, Czub, Majorek, Arolli, Variot, Anonick, Minor, Ballicora, Becker and Kuhn..
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.
| Medienart: |
E-Artikel |
|---|
| Erscheinungsjahr: |
2021 |
|---|---|
| Erschienen: |
2021 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:8 |
|---|---|
| Enthalten in: |
Frontiers in molecular biosciences - 8(2021) vom: 25., Seite 646046 |
| Sprache: |
Englisch |
|---|
| Beteiligte Personen: |
Baumgartner, Jackson T [VerfasserIn] |
|---|
| Links: |
|---|
| Themen: |
Acetylation |
|---|
| Anmerkungen: |
Date Revised 25.04.2022 published: Electronic-eCollection Citation Status PubMed-not-MEDLINE |
|---|
| doi: |
10.3389/fmolb.2021.646046 |
|---|
| funding: |
|
|---|---|
| Förderinstitution / Projekttitel: |
|
| PPN (Katalog-ID): |
NLM324658958 |
|---|
| LEADER | 01000naa a22002652 4500 | ||
|---|---|---|---|
| 001 | NLM324658958 | ||
| 003 | DE-627 | ||
| 005 | 20231225190805.0 | ||
| 007 | cr uuu---uuuuu | ||
| 008 | 231225s2021 xx |||||o 00| ||eng c | ||
| 024 | 7 | |a 10.3389/fmolb.2021.646046 |2 doi | |
| 028 | 5 | 2 | |a pubmed24n1082.xml |
| 035 | |a (DE-627)NLM324658958 | ||
| 035 | |a (NLM)33912589 | ||
| 040 | |a DE-627 |b ger |c DE-627 |e rakwb | ||
| 041 | |a eng | ||
| 100 | 1 | |a Baumgartner, Jackson T |e verfasserin |4 aut | |
| 245 | 1 | 0 | |a Gcn5-Related N-Acetyltransferases (GNATs) With a Catalytic Serine Residue Can Play Ping-Pong Too |
| 264 | 1 | |c 2021 | |
| 336 | |a Text |b txt |2 rdacontent | ||
| 337 | |a ƒaComputermedien |b c |2 rdamedia | ||
| 338 | |a ƒa Online-Ressource |b cr |2 rdacarrier | ||
| 500 | |a Date Revised 25.04.2022 | ||
| 500 | |a published: Electronic-eCollection | ||
| 500 | |a Citation Status PubMed-not-MEDLINE | ||
| 520 | |a Copyright © 2021 Baumgartner, Habeeb Mohammad, Czub, Majorek, Arolli, Variot, Anonick, Minor, Ballicora, Becker and Kuhn. | ||
| 520 | |a 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 | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a Gcn5-related N-acetyltransferase | |
| 650 | 4 | |a Pseudomonas aeruginosa | |
| 650 | 4 | |a acetylation | |
| 650 | 4 | |a acetyltransferase | |
| 650 | 4 | |a docking | |
| 650 | 4 | |a enzyme mechanism | |
| 650 | 4 | |a ping-pong kinetic mechanism | |
| 700 | 1 | |a Habeeb Mohammad, Thahani S |e verfasserin |4 aut | |
| 700 | 1 | |a Czub, Mateusz P |e verfasserin |4 aut | |
| 700 | 1 | |a Majorek, Karolina A |e verfasserin |4 aut | |
| 700 | 1 | |a Arolli, Xhulio |e verfasserin |4 aut | |
| 700 | 1 | |a Variot, Cillian |e verfasserin |4 aut | |
| 700 | 1 | |a Anonick, Madison |e verfasserin |4 aut | |
| 700 | 1 | |a Minor, Wladek |e verfasserin |4 aut | |
| 700 | 1 | |a Ballicora, Miguel A |e verfasserin |4 aut | |
| 700 | 1 | |a Becker, Daniel P |e verfasserin |4 aut | |
| 700 | 1 | |a Kuhn, Misty L |e verfasserin |4 aut | |
| 773 | 0 | 8 | |i Enthalten in |t Frontiers in molecular biosciences |d 2014 |g 8(2021) vom: 25., Seite 646046 |w (DE-627)NLM249149435 |x 2296-889X |7 nnns |
| 773 | 1 | 8 | |g volume:8 |g year:2021 |g day:25 |g pages:646046 |
| 856 | 4 | 0 | |u http://dx.doi.org/10.3389/fmolb.2021.646046 |3 Volltext |
| 912 | |a GBV_USEFLAG_A | ||
| 912 | |a GBV_NLM | ||
| 951 | |a AR | ||
| 952 | |d 8 |j 2021 |b 25 |h 646046 | ||