Abstract
C3′-deoxygenation of aminoglycosides results in their decreased susceptibility to phosphorylation thereby increasing their efficacy as antibiotics. However, the biosynthetic mechanism of C3′-deoxygenation is unknown. To address this issue, aprD4 and aprD3 genes from the apramycin gene cluster in Streptomyces tenebrarius were expressed in E. coli and the resulting gene products were characterized in vitro. AprD4 is shown to be a radical S-adenosylmethionine (SAM) enzyme, catalyzing homolysis of SAM to 5′-deoxyadenosine (5′-dAdo) in the presence of paromamine. [4′-2H]-Paromamine was prepared and used to show that its C4′-H is transferred to 5′-dAdo by AprD4, during which the substrate is dehydrated to a product consistent with 4′-oxolividamine. In contrast, paromamine is reduced to a deoxy product when incubated with AprD4/AprD3/NADPH. These results show that AprD4 is the first radical SAM diol-dehydratase and, along with AprD3, is responsible for 3′-deoxygenation in aminoglycoside biosynthesis.
| Original language | English |
|---|---|
| Pages (from-to) | 3724-3728 |
| Number of pages | 5 |
| Journal | Angewandte Chemie - International Edition |
| Volume | 55 |
| Issue number | 11 |
| DOIs | |
| Publication status | Published - 2016 Feb 15 |
| Externally published | Yes |
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Keywords
- aminoglycosides
- biosynthesis
- enzyme catalysis
- radical enzymes
- reaction mechanisms
ASJC Scopus subject areas
- Chemistry(all)
- Catalysis
Cite this
C3′-Deoxygenation of Paromamine Catalyzed by a Radical S-Adenosylmethionine Enzyme : Characterization of the Enzyme AprD4 and Its Reductase Partner AprD3. / Kim, Hak Joong; Levieux, Jake; Yeh, Yu Cheng; Liu, Hung Wen.
In: Angewandte Chemie - International Edition, Vol. 55, No. 11, 15.02.2016, p. 3724-3728.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - C3′-Deoxygenation of Paromamine Catalyzed by a Radical S-Adenosylmethionine Enzyme
T2 - Characterization of the Enzyme AprD4 and Its Reductase Partner AprD3
AU - Kim, Hak Joong
AU - Levieux, Jake
AU - Yeh, Yu Cheng
AU - Liu, Hung Wen
PY - 2016/2/15
Y1 - 2016/2/15
N2 - C3′-deoxygenation of aminoglycosides results in their decreased susceptibility to phosphorylation thereby increasing their efficacy as antibiotics. However, the biosynthetic mechanism of C3′-deoxygenation is unknown. To address this issue, aprD4 and aprD3 genes from the apramycin gene cluster in Streptomyces tenebrarius were expressed in E. coli and the resulting gene products were characterized in vitro. AprD4 is shown to be a radical S-adenosylmethionine (SAM) enzyme, catalyzing homolysis of SAM to 5′-deoxyadenosine (5′-dAdo) in the presence of paromamine. [4′-2H]-Paromamine was prepared and used to show that its C4′-H is transferred to 5′-dAdo by AprD4, during which the substrate is dehydrated to a product consistent with 4′-oxolividamine. In contrast, paromamine is reduced to a deoxy product when incubated with AprD4/AprD3/NADPH. These results show that AprD4 is the first radical SAM diol-dehydratase and, along with AprD3, is responsible for 3′-deoxygenation in aminoglycoside biosynthesis.
AB - C3′-deoxygenation of aminoglycosides results in their decreased susceptibility to phosphorylation thereby increasing their efficacy as antibiotics. However, the biosynthetic mechanism of C3′-deoxygenation is unknown. To address this issue, aprD4 and aprD3 genes from the apramycin gene cluster in Streptomyces tenebrarius were expressed in E. coli and the resulting gene products were characterized in vitro. AprD4 is shown to be a radical S-adenosylmethionine (SAM) enzyme, catalyzing homolysis of SAM to 5′-deoxyadenosine (5′-dAdo) in the presence of paromamine. [4′-2H]-Paromamine was prepared and used to show that its C4′-H is transferred to 5′-dAdo by AprD4, during which the substrate is dehydrated to a product consistent with 4′-oxolividamine. In contrast, paromamine is reduced to a deoxy product when incubated with AprD4/AprD3/NADPH. These results show that AprD4 is the first radical SAM diol-dehydratase and, along with AprD3, is responsible for 3′-deoxygenation in aminoglycoside biosynthesis.
KW - aminoglycosides
KW - biosynthesis
KW - enzyme catalysis
KW - radical enzymes
KW - reaction mechanisms
UR - http://www.scopus.com/inward/record.url?scp=84976240409&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84976240409&partnerID=8YFLogxK
U2 - 10.1002/anie.201510635
DO - 10.1002/anie.201510635
M3 - Article
VL - 55
SP - 3724
EP - 3728
JO - Angewandte Chemie - International Edition
JF - Angewandte Chemie - International Edition
SN - 1433-7851
IS - 11
ER -