Valeria Di Giacomo, Senior R&D Researcher in ZeClinics, presents a new paper in collaboration with Qatar University.
Hesham M Ismail 1 2, Navaneethakrishnan Krishnamoorthy 3 4, Nader Al-Dewik 5, Hatem Zayed 1, Nura A Mohamed 2, Valeria Di Giacomo 6, Sapna Gupta 7, Johannes Häberle 8, Beat Thöny 8, Henk J Blom 9, Waren D Kruger 7, Tawfeg Ben-Omran 5, Gheyath K Nasrallah 1 2
1Department of Biomedical Sciences, College of Health Sciences, Qatar University, Doha, Qatar.
2Biomedical Research Center, Qatar University, Doha, Qatar.
3Systems Biology Department, Sidra Medical and Research Centre, Doha, Qatar.
4Heart Science Centre, National Heart and Lung Institute, Imperial College London, London, UK.
5Clinical and Metabolic Genetics, Department of Pediatrics, Hamad Medical Corporation, Doha, Qatar.
6ZeClinics SL, PRBB (Barcelona Biomedical Research Park), Barcelona, Spain.
7Cancer Biology Program, Fox Chase Cancer Center, Philadelphia, Pennsylvania.
8University Children's Hospital and Children's Research Center, Division of Metabolism, Zurich, Switzerland.
9Laboratory of Clinical Biochemistry and Metabolism, Department of General Pediatrics, Adolescent Medicine and Neonatology, University Medical Centre Freiburg, Freiburg, Germany.
ABSTRACT
Homocystinuria is a rare inborn error of methionine metabolism caused by cystathionine β-synthase (CBS) deficiency. The prevalence of homocystinuria in Qatar is 1:1,800 births, mainly due to a founder Qatari missense mutation, c.1006C>T; p.R336C (p.Arg336Cys). We characterized the structure-function relationship of the p.R336C-mutant protein and investigated the effect of different chemical chaperones to restore p.R336C-CBS activity using three models: in silico, ΔCBS yeast, and CRISPR/Cas9 p.R336C knock-in HEK293T and HepG2 cell lines. Protein modeling suggested that the p.R336C induces severe conformational and structural changes, perhaps influencing CBS activity. Wild-type CBS, but not the p.R336C mutant, was able to restore the yeast growth in ΔCBS-deficient yeast in a complementation assay. The p.R336C knock-in HEK293T and HepG2 cells decreased the level of CBS expression and reduced its structural stability; however, treatment of the p.R336C knock-in HEK293T cells with betaine, a chemical chaperone, restored the stability and tetrameric conformation of CBS, but not its activity. Collectively, these results indicate that the p.R336C mutation has a deleterious effect on CBS structure, stability, and activity, and using the chemical chaperones approach for treatment could be ineffective in restoring p.R336C CBS activity.
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