Journal
JOURNAL OF BIOLOGICAL CHEMISTRY
Volume 298, Issue 8, Pages -Publisher
ELSEVIER
DOI: 10.1016/j.jbc.2022.102166
Keywords
-
Categories
Funding
- NSF [1654611]
- National Cancer Institute [ACB-12002]
- NIH [S10 OD012289, DE-AC02-76SF00515]
- National Institute of General Medical Sciences [AGM-12006, P30GM138396]
- Argonne National Laboratory [DE-AC02-06CH11357]
- U.S. Department of Energy, Office of Science, Office of Basic Energy Sciences [P30GM133894]
- DOE Office of Biological and Environmental Research
- National Institutes of Health, National Institute of General Medical Sciences
- [R35GM136235]
- Direct For Mathematical & Physical Scien
- Division Of Chemistry [1654611] Funding Source: National Science Foundation
Ask authors/readers for more resources
Siderophores are metabolites used by microbes to sequester environmental iron. Streptomyces strains produce desferrioxamine (DFO) siderophores, which consist of repeating units of hydroxamate-based desferrioxamine and succinate. In this study, we characterized four DesD orthologs from Streptomyces strains that produce unique DFO siderophores. We synthesized a structural mimic of N-1-hydroxy-N-1-succinyl-cadaverine and obtained crystal structures of DesD in different forms. Our findings suggest that acyl-AMS derivatives can be useful tools to study the mechanism and structure of this family of enzymes.
Siderophores are conditionally essential metabolites used by microbes for environmental iron sequestration. Most Streptomyces strains produce hydroxamate-based desferriox-amine (DFO) siderophores composed of repeating units of N-1- hydroxy-cadaverine (or N-1-hydroxy-putrescine) and succinate. The DFO biosynthetic operon, desABCD, is highly conserved in Streptomyces; however, expression of desABCD alone does not account for the vast structural diversity within this natural product class. Here, we report the in vitro reconstitution and biochemical characterization of four DesD orthologs from Streptomyces strains that produce unique DFO siderophores. Under in vitro conditions, all four DesD orthologs displayed similar saturation steady-state kinetics (V-max = 0.9-2.5 mu M.min(-1)) and produced the macrocyclic trimer DFOE as the favored product, suggesting a conserved role for DesD in the biosynthesis of DFO siderophores. We further synthesized a structural mimic of N-1-hydroxy-N-1- succinyl-cadaverine (HSC)-acyl-adenylate, the HSC-acyl sulfamoyl adenosine analog (HSC-AMS), and obtained crystal structures of DesD in the ATP-bound, AMP/PPi-bound, and HSC-AMS/Pi-bound forms. We found HSC-AMS inhibited DesD orthologs (IC50 values = 48-53 mu M) leading to accumulation of linear trimeric DFOG and di-HSC at the expense of macrocyclic DFOE. Addition of exogenous PPi enhanced DesD inhibition by HSC-AMS, presumably via stabilization of the DesD-HSC-AMS complex, similar to the proposed mode of adenylate stabilization where PPi remains buried in the active site. In conclusion, our data suggest that acyl-AMS derivatives may have utility as chemical probes and bisubstrate inhibitors to reveal valuable mechanistic and structural insight for this unique family of adenylating enzymes.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available