Identification of redox-active cell-surface proteins by mechanism-based kinetic trapping

Area of Science:

• Biochemistry
• Cell Biology
• Proteomics

Background:

• Extracellular thiol-dependent oxidoreductases influence cell-surface processes like integrin activation and viral fusion.
• Tumor cells often upregulate these oxidoreductases, suggesting roles in tumor growth and microenvironment remodeling.
• Identifying specific cell-surface targets of these enzymes has been challenging due to the transient nature of thiol-disulfide exchange.

Purpose of the Study:

• To develop and apply a novel activity-based proteomics approach for identifying cell-surface proteins engaged in disulfide exchange.
• To overcome limitations of conventional methods in detecting transient thiol-disulfide reactions.

Main Methods:

• Utilized mechanism-based kinetic trapping, a form of activity-based proteomics.
• Applied the method to study interactions involving thioredoxin-1.

Main Results:

• Successfully identified individual cell-surface target proteins involved in disulfide exchange with thiol-dependent oxidoreductases.
• Demonstrated the utility of the kinetic trapping approach for studying extracellular redox enzyme targets.

Conclusions:

• Mechanism-based kinetic trapping is an effective strategy for identifying cell-surface targets of extracellular redox enzymes.
• The approach is adaptable for studying other members of the thioredoxin superfamily and similar redox enzymes.