Related Experiment Videos
The Toxoplasma adhesive protein MIC2 is proteolytically processed at multiple sites by two parasite-derived proteases
V B Carruthers1, G D Sherman, L D Sibley
1Department of Molecular Microbiology, Washington University School of Medicine, St. Louis, Missouri 63110, USA. vcarruth@jhsph.edu
Abstract:
MIC2 is an adhesive protein that participates in host cell invasion by the obligate intracellular parasite Toxoplasma gondii. Earlier studies established that MIC2 is secreted into the culture medium by extracellular parasites and that release is coincident with proteolytic modification. Since little is known about proteolytic processing of proteins secreted by T. gondii, we undertook this study to investigate the proteolytic events that accompany secretion of MIC2. We demonstrate that the C-terminal domain of MIC2 is removed by a protease, termed MPP1, when MIC2 is released into the culture supernatant. Additionally, prior to release, a second protease, termed MPP2, trims the N terminus of MIC2, resulting in the release of heterogeneously sized species of MIC2. Although MPP1 activity was unaffected by any of the protease inhibitors tested, MPP2 activity was blocked by a subset of serine and cysteine protease inhibitors. These results establish that MIC2 is proteolytically modified at multiple sites by two distinct enzymes that probably operate on the parasite surface.
Insights
Two proteases, MIC2-associated proteinase 1 (MPP1) and MPP2, modify the MIC2 protein during secretion by Toxoplasma gondii. These enzymes process the protein at different sites, impacting parasite invasion mechanisms.
Area of Science:
- Parasitology
- Molecular Biology
- Biochemistry
Background:
- MIC2 is a key adhesive protein facilitating host cell invasion by Toxoplasma gondii.
- Secretion of MIC2 by extracellular parasites is linked to proteolytic modification.
- Limited knowledge exists regarding the proteolytic processing of secreted T. gondii proteins.
Purpose of the Study:
- To investigate the specific proteolytic events involved in MIC2 secretion.
- To identify the proteases responsible for MIC2 processing.
- To understand the functional implications of MIC2 modification during invasion.
Main Methods:
- Protease activity assays using protease inhibitors.
- Analysis of MIC2 processing products in culture supernatant.
- N-terminal and C-terminal domain analysis of secreted MIC2.
Main Results:
- A C-terminal domain of MIC2 is removed by MIC2-associated proteinase 1 (MPP1) during secretion.
- A second protease, MPP2, trims the N terminus of MIC2 before release, creating varied protein sizes.
- MPP1 activity is resistant to tested inhibitors, while MPP2 is inhibited by serine and cysteine protease inhibitors.
Conclusions:
- MIC2 undergoes multi-site proteolytic modification by two distinct proteases, MPP1 and MPP2.
- These processing events likely occur on the parasite surface.
- Understanding MIC2 processing provides insights into Toxoplasma gondii invasion strategies.