title: The application of advanced structural bioinformatics methods to parasitic membrane proteins
Creators: name: Murphy, David
Publisher: name: University of Liverpool Repository
Year: 2024

description: Malaria and toxoplasmosis are globally burdensome diseases caused by the still poorly understood parasites Plasmodium falciparum and Toxoplasma gondii belonging to the Apicomplexa phylum. This phylum is highly divergent to most known eukaryotes meaning that many protein sequences are beyond recognition by homology-based methods, and a great proportion of the proteins in these parasites lack any known domains and functions. Part of the parasitic success of Apicomplexans is due to the presence of the biosynthetic powerhouse that is the apicoplast organelle, with the ATP-Synthase complex also being critical to the parasites. Though of course, other parasitic membrane contexts are also worthy of attention. The apicoplast additionally presents experimental difficulties; hence illuminating more putative molecular functions of the organelle’s uncharacterised proteome via alternative in silico approaches would greatly aid in the understanding of its molecular biology further, and may even reveal new drug targets.

In recent years there have been strides in the ability to accurately predict contacts and distances of intra-protein residues from deep machine learning which have been exploited to improve our ability to accurately predict the structure of proteins, with one important example being AlphaFold2. Accurate structure models can help gauge the approximate molecular functions of proteins, hence modelling parasitic proteins of unknown structure and function could aid in understanding their function. Membrane proteins are a protein subtype which are much less well represented in the PDB due to various experimental difficulties. This makes parasitic membrane proteins of unknown structure and function a promising research niche for the application of advanced structural bioinformatics methods.

This study focuses on the prediction of structure to infer the function of 19 transmembrane proteins. Our primary focus corresponded to 14 of these which were either experimentally localised or predicted to be targeted to the P. falciparum apicoplast, though at least five here possessed alternative localisations, including export to the host erythrocyte. Based primarily on the presence of fold matches to our AlphaFold2 models, the results of these analyses infer some of these proteins to possess protein binding and transporter functions, though new insights were also revealed for some of the other proteins. One of the most crucial findings was that the apicoplast membrane protein PF3D7_0622700 contains a single copy of the UVB_Sens_prot domain which escapes sequence-based detection due mostly to the presence of a particularly large loop region between the fourth and fifth transmembrane alpha-helices. The presence of the domain and its predicted structure renders it a member of the major facilitator superfamily of transporters, and based on both literature and our results, could hypothetically transport the active form of vitamin B6 into the apicoplast. Our other research sub-project to predict the structure of five uncharacterised subunits from the T. gondii ATP-Synthase complex was superseded by the unexpected release of the complex’s experimental structure. Centred on this structure we discuss why structure modelling was unsuccessful for this particular type of membrane protein.