Assays

Prepared multimeric tubulin protein receptors for docking studies, generated by alignment of two identical models of alpha-tubulin on alpha-tubulin chains of two neighboring protofilaments.

Docking results of trifluraline and dinitroaniline-etherphospholipid hybrids against different kinetoplastid alpha-tubulin receptors with an induced fit docking routine. The docking protocol involves an initial crude ligand placement step, subsequent receptor optimization in response to ligand binding, and another docking step into the optimized receptor.

Creation of homology models of various tubulins from dinitroaniline-sensitive and -resistant species, and a comparative analysis of their electrostatic potential grids overall and in putative binding site regions using PIPSA (Protein Interaction Property Similarity Analysis).

Alignments of various alpha-tubulin and beta-tubulin sequences from dinitroaniline-sensitive and dinitroaniline-resistant species. Sequences were retrieved from UniProt with the identifiers listed below and subjected to a multiple sequence alignment using ClustalOmega (ebi.ac.uk/Tools/msa/clustalo/; ClustalOmega webserver, last accessed 16-02-23):

alpha-tubulin:

• [dinitroaniline sensitive] T. cruzi - Q27352; T. brucei brucei - Q4GYY5; L. infantum - ...

Incrementally increase the activity of some target rescue enzymes from 20% of default expression to 200% of default expression in a control and MCADD model to see if flux and CoASH concentration are rescued. Generates Fig. 6, S5, and S6.

Download "Model_notebooks.rar", unzip, and run "11, Fig6+S5-rescues-[needs-(1-and-10)]-20221109.nb", "10, Fig6B-inset-rescues-(low-acetylCoA)-[needs-(1)]-20221109.nb", and "16, FigS6-rescues-20221109-fixed-[needs-(1)]-CoASH.nb" after running "1, generate-model-20221109.nb" ...

Based on odendaal1, a control model is made and compared to model deficient for short-chain acyl-CoA dehydrogenase (SCADD, 0%), medium-chain acyl-CoA dehydrogenase (MCADD, 0%), and very long-chain acyl-CoA dehydrogenase (VLCADD, 10%). With and withou metabolite partitioning, and with a fixed mitohondrial free CoASH. Generates Figures 3, 4, S1, S2, and S3 in the related paper.

Download "Model_notebooks.rar", unzip, and run: "7, Fig3+4+S1+S3-ACADDs-[needs-(1)]-20221109.nb" and "14, ...