In the Derbyshire Lab, my project focuses on studying the interaction between a Plasmodium falciparum parasite protein, PfHsp70-1 and a phospholipid, PI3P.
Plasmodium falciparum is a unicellular, protozoan, parasite species that infects humans. It is transmitted though mosquito bites and results in falciparum malaria—the most malignant form of this disease. When a bite occurs, sporozoites (forms of Plasmodium falciparum that result from a division of the cyst containing the parasite zygotes) that have clustered in a mosquito’s salivary glands enter into the bloodstream. From there, they enter and infect liver cells and multiply, rupturing to produce merozoites. Merozoites invade and infect red blood cells initiating the symptoms of malaria.
Heat shock proteins within P. falciparum act as molecular chaperones, helping in the folding/unfolding of other macromolecules. The Hsp70 family is believed to be essential for the life cycle of this parasite, as proteins in the family help mitigate the effects of changing temperature (and other physiological stressors) when the parasite is passing from the cold-blooded mosquito to the warm-blooded human host. It also facilitates invasion of host cells (by potentially trafficking parasite proteins), and increases heat resistance/resilience in the blood stage.
Within the Hsp70 family is PfHsp70-1, a protein mainly found in the cytosol of the parasite cell. PfHsp70-1 is expressed in the blood stages of malaria and is believed to be a potential exporter of parasite proteins into red blood cells during infection. It has been proposed that PfHsp70-1 works with Hsp90 for parasite growth and development, possibly helps regulate the parasite’s protein translation, and trafficking proteins into the apicoplast, a parasite organelle. The linker interface on PfHsp70-1 is believed to be responsible for communication and interaction with Hsp40 for parasite development. Due to these properties, PfHSp70-1 has been studied closely in vaccine research.
PI(3)P is a phospholipid that regulates cellular functions and assists in vesicular trafficking of proteins. Plasmodium PI3-kinase has been shown to be essential for P. falciparum growth. During red blood cell infection, it is believed that P. falciparum forms a protein and lipid trafficking system by potentially synthesizing PI(3)P. According to Mbengue et al, PI3P works to transport parasite proteins from the Plasmodium endoplasmic reticulum, into the red blood cell during blood-stage infection. However, not much is known about the actual mechanism for trafficking, thus making it a target for vaccine research and development. It is important to study PI3P as it is a key component in the blood stage of falciparum malaria.
The Derbyshire lab has previously identified that the C-terminal domain of PfHsp70-1 may mediate its interaction with PI3P, which could play an important role in parasite survival. My project deals with the potential interaction between PfHsp70-1 and PI(3)P and the part of the structure involved with binding between the two. I will be determining the binding quantitatively via isothermal titration calorimetry.
So far, I have worked to induce protein expression (developed via yeast cell culture) and protein purification of Hsp70-1 WT. I am currently being trained on ITC so fingers crossed that I will be able to get some data on the binding affinity!
Lab has been an amazing experience these past two weeks, and I’ve been learning a lot under the guidance of some pretty great teachers. I’m looking forward to learning more and diving deeper into this project!
