In silico and in vivo models for Qatari-specific classical homocystinuria as basis for development of novel therapies

Hesham M. Ismail, Navaneethakrishnan Krishnamoorthy, Nader Al-Dewik, Hatem Zayed, Nura A. Mohamed, Valeria Di Giacomo, Sapna Gupta, Johannes Häberle, Beat Thöny, Henk J. Blom, Waren D. Kruger, Tawfeg Ben-Omran, Gheyath K. Nasrallah

Research output: Contribution to journalArticle

1 Citation (Scopus)

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.

Original languageEnglish
JournalHuman Mutation
DOIs
Publication statusAccepted/In press - 1 Jan 2018

Fingerprint

Cystathionine
Homocystinuria
Computer Simulation
Therapeutics
Yeasts
Hep G2 Cells
Clustered Regularly Interspaced Short Palindromic Repeats
Qatar
Inborn Errors Metabolism
Betaine
Missense Mutation
Mutant Proteins
Methionine

Keywords

  • CBS
  • chemical chaperones
  • Homocystinuria
  • in silico
  • in vivo models
  • p.R336C mutation
  • Qatar

ASJC Scopus subject areas

  • Genetics
  • Genetics(clinical)

Cite this

In silico and in vivo models for Qatari-specific classical homocystinuria as basis for development of novel therapies. / Ismail, Hesham M.; Krishnamoorthy, Navaneethakrishnan; Al-Dewik, Nader; Zayed, Hatem; Mohamed, Nura A.; Giacomo, Valeria Di; Gupta, Sapna; Häberle, Johannes; Thöny, Beat; Blom, Henk J.; Kruger, Waren D.; Ben-Omran, Tawfeg; Nasrallah, Gheyath K.

In: Human Mutation, 01.01.2018.

Research output: Contribution to journalArticle

Ismail, HM, Krishnamoorthy, N, Al-Dewik, N, Zayed, H, Mohamed, NA, Giacomo, VD, Gupta, S, Häberle, J, Thöny, B, Blom, HJ, Kruger, WD, Ben-Omran, T & Nasrallah, GK 2018, 'In silico and in vivo models for Qatari-specific classical homocystinuria as basis for development of novel therapies', Human Mutation. https://doi.org/10.1002/humu.23682
Ismail, Hesham M. ; Krishnamoorthy, Navaneethakrishnan ; Al-Dewik, Nader ; Zayed, Hatem ; Mohamed, Nura A. ; Giacomo, Valeria Di ; Gupta, Sapna ; Häberle, Johannes ; Thöny, Beat ; Blom, Henk J. ; Kruger, Waren D. ; Ben-Omran, Tawfeg ; Nasrallah, Gheyath K. / In silico and in vivo models for Qatari-specific classical homocystinuria as basis for development of novel therapies. In: Human Mutation. 2018.
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AU - Zayed, Hatem

AU - Mohamed, Nura A.

AU - Giacomo, Valeria Di

AU - Gupta, Sapna

AU - Häberle, Johannes

AU - Thöny, Beat

AU - Blom, Henk J.

AU - Kruger, Waren D.

AU - Ben-Omran, Tawfeg

AU - Nasrallah, Gheyath K.

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N2 - 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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