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C ritical Assessment of Genome Interpretation

Frataxin

Challenge: Fraxatin

Variant data: public

Last updated: 14 April 2018

This challenge is closed.

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Summary

Frataxin is a highly conserved protein found in prokaryotes and eukaryotes that is required for efficient regulation of cellular iron homeostasis. Humans with a frataxin deficiency have the cardio- and neurodegenerative disorder Friedreich’s Ataxia. A library of eight missense variants was assessed by near and far-UV circular dichroism and intrinsic fluorescence spectra to determine thermodynamic stability at different concentration of denaturant. These data were used to calculate a ΔΔG_H20 value, the difference in unfolding free energy ΔΔG_H20 between the variant and wildtype proteins for each variant. The challenge is to predict ΔΔG_H20 for each frataxin variant.

Background

The human frataxin is a protein localized in the mitochondrion and cytoplasm that promotes the heme biosynthesis and assembly and repair of iron-sulphur clusters by delivering Fe²⁺ to proteins involved in these pathways. The protein may play a role in protection against iron-catalyzed oxidative stress through its ability to catalyze the oxidation of Fe²⁺ to Fe³⁺. The oligomeric form but not the monomeric form of the protein has ferroxidase activity in vitro. Oligomerization may provide a mechanism by which the protein stores large amounts of iron in the form of a ferrihydrite mineral.

Reduced expression of frataxin is the cause of Friedreich's Ataxia (FRDA), a lethal neurodegenerative disease. According to SwissVar database, eight single amino acids variants (p.Leu106Ser, p.Asp122Tyr, p.Gly130Val, p.Ile154Phe, p.Trp155Arg, p.Arg165Cys, p.Leu182Phe and p.Leu198Arg) have been associated to FRDA (Corey, 2016). The role of frataxin in cancer is still ambiguous: studies have shown that frataxin protects tumor cells against oxidative stress and apoptosis, but also acts as a tumor suppressor (Schulz et al., 2006; Guccini et al., 2011).

The single amino acid variants included in the Frataxin challenge were selected from the COSMIC (Catalog of Somatic Mutations in Cancer) database. These are somatic variants associated with neoplastic diseases and/or detected in cancer tissues. From the 21 variants reported in COSMIC we included in this challenge those that could be obtained as a soluble recombinant protein in E. coli, and excluded the variants whose expression failed.

The results of previous thermodynamic experiments on the wildtype and a different set of mutated human frataxin have been reported (Faraj et al., 2016).

Experiment

Human frataxin variants are obtained with specific mutagenesis primers by PCR, using wild-type protein as a template. Wild-type and mutants are then expressed in E. coli and purified. The structural conformation of the variants is compared to that of the wild-type by monitoring the near and far-UV circular dichroism and intrinsic fluorescence spectra. The thermodynamic stability is measured at a different concentration of denaturant (Urea) by monitoring the spectral changes (far-UV circular dichroism and intrinsic fluorescence emission) induced by urea. The spectral changes are fitted to zero denaturant concentration (ΔG_H20).

Prediction challenge

For each variant, participants are asked to predict the ΔΔG_H20 value, which is the difference in unfolding free energy (ΔG_H20) between the mutant and wild-type proteins, in kcal/mol. The predictions will be evaluated via several methods including those that consider the difference between the predicted and experimental ΔΔG_H20 values. In addition, predictions will be evaluated by dividing the variants into two classes: the significantly destabilizing mutants with ΔΔG_H20 ≤ -1.0 kcal/mol, and the neutral/stabilizing mutants with ΔΔG_H20 > -1.0 kcal/mol..

Prediction submission format

The prediction submission is a tab-delimited text file. Organizers provide a template file, which must be used for submission. In addition, a validation script is provided, and predictors must check the correctness of the format before submitting their predictions. Each data row in the submitted file must include the following columns:

AA substitution - Single-letter amino acid substitution and position; e.g., D104G (relative to UniProt protein Q16595 (FRDA_HUMAN).
Value -ΔΔG_H20 value
Standard deviation - SD of the prediction in column 2.
Comment – optional brief comment on the basis of the prediction in column 2.

In the template file, cells in columns 2-4 are marked with a "*". Submit your predictions by replacing the "*" with your value. No empty cells are allowed in the submission. For a given subset, you must submit predictions and standard deviations for all or none of the variants; if you are not confident in a prediction for a variant, enter an appropriate large standard deviation for the prediction. Optionally, enter a brief comment on the basis of the prediction. If you do not enter a comment on a prediction, leave the "*" in those cells. Please make sure you follow the submission guidelines strictly.

In addition, your submission should include a detailed description of the method used to make the predictions, similar to the style of the Methods section in a scientific article. This information must be submitted as a separate file.

To submit predictions, you need to create or be part of a CAGI User group. Submit your predictions by accessing the link "All submission forms" from the front page of your group. For more details, please read the FAQ page.

Additional information

The function of the human Frataxin and other information are reported in UniProt and are accessible with the identifier Q16595 (FRDA_HUMAN).

The structure of human Frataxin from position 90 to 208 has been determined by X-ray crystallography and is available from the PDB with code 1EKG.

References

Corey DR. Synthetic nucleic acids and treatment of neurological diseases. JAMA Neurol (2016) 73(10):1238-1242. PubMed

Faraj SE, et al. Human frataxin folds via an intermediate state. Role of the c-terminal region. Sci Rep (2016) 6:20782. PubMed

Guccini I, et al. Frataxin participates to the hypoxia-induced response in tumors. Cell Death Dis (2014) 2(2):e123. PubMed

Schulz TJ, et al. Induction of oxidative metabolism by mitochondrial frataxin inhibits cancer growth: Otto Warburg revisited. J Biol Chem (2006) 281(2):977-981. PubMed

Assessment

This challenge will be assessed by Emidio Capriotti, University of Bologna, Italy.

Download dataset: Fraxatin_dataset.txt (3.5 KB)

Download submission template: Fraxatin_submission_template.txt

Download submission validation script: Validation Perl script (not available)

Dataset providers

The dataset is provided by the group of Roberta Chiaraluce and Valerio Consalvi, Sapienza University, Roma, Italy.