Huntington’s disease is caused by a gene mutation that causes an abnormal form of the protein huntingtin to build up in the brain. Small chemical modifications on different parts of huntingtin could reduce its toxicity and aggregation, but as many enzymes already chemically modify the protein in the cell, it has been difficult to determine what chemical modifications could serve as future therapies.
In a world first, scientists have now developed synthetic methods that allow site-specific chemical modifications on huntingtin while bypassing the need to identify the enzymes behind them.
The study was published in Angewandte Chemie.
After being produced by its gene, the huntingtin protein is subjected to numerous chemical changes, during which enzymes in the cell attach different chemical groups to it, such as phosphate (phosphorylation) or acetylene (acetylation). These changes are called “post-translational modifications” but the key molecular players that regulate them remain unknown.
To address this knowledge gap and explore the therapeutic potential of post-translational modifications, a group of researchers turned to chemistry and developed synthetic strategies that allow site-specific introduction of chemical modifications. The new strategy combines chemical and bacterial synthesis of proteins to generate a part of mutant huntingtin where many important post-translational modifications take place. This segment is known as “mutant hutingtin exon1” (Httex1) and has been shown to be sufficient for reproducing key features of Huntington’s disease in animal models.
Using the method, the researchers could now generate all the known modified forms of huntingtin in very pure and homogeneous forms.
The researchers explored how these modifications can act as molecular switches that can be exploited to regulate Huntingtin structure, function and toxicity. In this vein, they made three discoveries regarding the relationship between post-translational modifications of Huntingtin and its structure.
First, phosphorylation of threonine on position 3 (T3) stabilizes a helical formation of huntingtin’s first 17 amino acids, and interferes with its ability to aggregate. This could explain why this particular modification is reduced in Huntington’s disease and suggest that modulating the level of this modification could protect against HD disease.
Second, the study found that substituting threonine with another amino acid (glutamate or aspartate) to mimic the charge of the phosphate group, does not fully reproduce the effects of genuine phosphorylation on the structure and aggregation of huntingtin. This is important for biologists who, in the absence of knowledge about the key enzymes that regulate phosphorylation, use this substitution as a go-to solution.
Finally, the scientists were able for the first time to study the impact of acetylation, which takes place on three Lysine amino acids of huntingtin. While acetylation of individual residues did affect huntingtin’s structure or aggregation, acetylation at one lysine (in position 6) did reverse the protective effect of T3 phosphorylation when the two modifications were introduced simultaneously.
The researchers say that their study highlights the importance of better understanding the enzymes that regulate huntingtin modifications and note that their research could lead to the establishment of new biomarkers for disease diagnosis and monitoring.
Paper: “Mutant Exon1 Huntingtin Aggregation is Regulated by T3 Phosphorylation-Induced Structural Changes and Crosstalk between T3 Phosphorylation and Acetylation at K6”
Reprinted from materials provided by Ecole Polytechnique Fédérale de Lausanne.