Combination of SAXS and Protein Painting Discloses the Three-Dimensional Organization of the Bacterial Cysteine Synthase Complex, a Potential Target for Enhancers of Antibiotic Action.
Brenda RosaMarialaura MarchettiGianluca ParediHeinz AmenitschNina FrankoRoberto BenoniBarbara GiabbaiMaria Giovanna De MarinoAndrea MozzarelliLuca RondaPaola StoriciBarbara CampaniniStefano BettatiPublished in: International journal of molecular sciences (2019)
The formation of multienzymatic complexes allows for the fine tuning of many aspects of enzymatic functions, such as efficiency, localization, stability, and moonlighting. Here, we investigated, in solution, the structure of bacterial cysteine synthase (CS) complex. CS is formed by serine acetyltransferase (CysE) and O-acetylserine sulfhydrylase isozyme A (CysK), the enzymes that catalyze the last two steps of cysteine biosynthesis in bacteria. CysK and CysE have been proposed as potential targets for antibiotics, since cysteine and related metabolites are intimately linked to protection of bacterial cells against redox damage and to antibiotic resistance. We applied a combined approach of small-angle X-ray scattering (SAXS) spectroscopy and protein painting to obtain a model for the solution structure of CS. Protein painting allowed the identification of protein-protein interaction hotspots that were then used as constrains to model the CS quaternary assembly inside the SAXS envelope. We demonstrate that the active site entrance of CysK is involved in complex formation, as suggested by site-directed mutagenesis and functional studies. Furthermore, complex formation involves a conformational change in one CysK subunit that is likely transmitted through the dimer interface to the other subunit, with a regulatory effect. Finally, SAXS data indicate that only one active site of CysK is involved in direct interaction with CysE and unambiguously unveil the quaternary arrangement of CS.
Keyphrases
- protein protein
- small molecule
- high resolution
- fluorescent probe
- living cells
- crispr cas
- single molecule
- induced apoptosis
- oxidative stress
- binding protein
- transcription factor
- amino acid
- protein kinase
- air pollution
- hydrogen peroxide
- electronic health record
- molecular dynamics
- ms ms
- nitric oxide
- human health
- atomic force microscopy
- deep learning
- magnetic resonance
- big data
- risk assessment
- molecular dynamics simulations
- drug induced
- solid state
- climate change
- dual energy
- case control