On the function of TRAP substrate-binding proteins: the isethionate-specific binding protein IseP

Area of Science:

• Microbiology
• Structural Biology
• Biochemistry

Background:

• Bacteria utilize specialized transporters to acquire essential nutrients and adapt to diverse environments.
• The import of isethionate by sulfate-reducing bacteria is relevant to human health and industrial processes.
• Tripartite ATP-independent periplasmic (TRAP) transporters are crucial for nutrient uptake in many microorganisms.

Purpose of the Study:

• To elucidate the molecular mechanism of isethionate import by the TRAP transporter OaIsePQM in Oleidesulfovibrio alaskensis.
• To characterize the substrate-binding protein OaIseP and its interaction with isethionate.
• To provide structural insights into TRAP transporter function with sulfonate-containing substrates.

Main Methods:

• Isothermal titration calorimetry (ITC) to determine isethionate binding affinity to OaIseP.
• X-ray crystallography to obtain structures of OaIseP in ligand-free and isethionate-bound states.
• Analysis of protein structures to understand domain closure and ligand binding.

Main Results:

• The binding affinity of isethionate to OaIseP was determined as KD = 0.95 µM.
• X-ray structures revealed that OaIseP adopts a closed conformation upon isethionate binding.
• Sulfonate-containing buffer components (HEPES, MES) bound to the active site but did not induce domain closure, likely due to their larger size.

Conclusions:

• The study provides detailed molecular insights into how TRAP transporters bind sulfonate-containing substrates like isethionate.
• The findings highlight differences in ligand binding compared to TRAP transporters that bind carboxylate-containing substrates.
• This research may inform the development of novel antibiotics targeting TRAP transporters and guide protein engineering efforts.