Analyzing the Effects of a G137V Mutation in the FXN Gene.

Nathalie Faggianelli,Tommaso Vannocci,Roberto Testi,Silvia Romano,Rita Puglisi,Marina Frontali,Silvia Fortuni,Annalisa Pastore,Liana Veneziano

doi:10.3389/fnmol.2015.00066

Abstract

Reduced levels of frataxin, an essential mitochondrial protein involved in the regulation of iron-sulfur cluster biogenesis, are responsible for the recessive neurodegenerative Friedreich Ataxia (FRDA). Expansion of a GAA triplet in the first intron of the FRDA is essential for disease development which causes partial silencing of frataxin. In the vast majority of cases, patients are homozygotes for the expansion, but a small number of FRDA patients are heterozygotes for expansion and point mutations in the frataxin coding frame. In this study, we analyze the effects of a point mutation G137V. The patient P94–2, with a history of alcohol and drug abuse, showed a FRDA onset at the border between the classic and late onset phenotype. We applied a combination of biophysical and biochemical methods to characterize its effects on the structure, folding and activity of frataxin. Our study reveals no impairment of the structure or activity of the protein but a reduced folding stability. We suggest that the mutation causes misfolding of the native chain with consequent reduction of the protein concentration in the patient and discuss the possible mechanism of disease.

Highlights

Friedreich ataxia (FRDA, OMIM 229300) is the most common form of recessively inherited ataxia, a neurodegenerative disease associated to loss of voluntary movement (Pastore and Puccio, 2013)
We explored the structure-function relationship and carried out a detailed investigation of the possible consequences of this mutation on the frataxin fold, stability and function using complementary biochemical and biophysical structural techniques in vitro which include Circular Dichroism (CD), absorbance and Nuclear Magnetic Resonance (NMR) spectroscopies and enzymatic assays
When we compared the effect of the G130V mutant in the assay with that of wild-type frataxin, we did not observe any marked difference of behavior comparing the effect of wild-type frataxin and the mutant (Figure 5) suggesting that mutation has no a direct functional effect atleast under the conditions of this assay. These results indicate that, while affecting the stability of the protein, the G137V mutation does not significantly impair function, that is, when frataxin is folded, its function is not affected by the mutation

Summary

Introduction

Friedreich ataxia (FRDA, OMIM 229300) is the most common form of recessively inherited ataxia, a neurodegenerative disease associated to loss of voluntary movement (Pastore and Puccio, 2013). The severity of the disease is correlated with the number of repeats (Pastore and Puccio, 2013), which is lower than 30 in normal alleles and between 66 and 1700 in FRDA patients (Clark et al, 2004). FRDA patients have frataxin levels in peripheral tissues that range from 2–30% as compared to control levels (Deutsch et al, 2010; Nachbauer et al, 2011; Saccà et al, 2011, 2013). These levels correlate directly with age of onset and inversely with the number of GAA repeats. A certain variability is observed which makes it difficult to correlate precisely the frataxin levels with the severity of phenotype

Methods

Results

Conclusion