Modelling hCDKL5 Heterologous Expression in Bacteria
hCDKL5 refers to the human cyclin-dependent kinase like 5 that is primarily expressed in the brain. Mutations in its coding sequence are often causative of hCDKL5 deficiency disorder, a devastating neurodevelopmental disorder currently lacking a cure. The large-scale recombinant production of hCDKL5...
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MDPI AG
2021-07-01
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| Series: | Metabolites |
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| Online Access: | https://www.mdpi.com/2218-1989/11/8/491 |
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| author | Marco Fondi Stefano Gonzi Mikolaj Dziurzynski Paola Turano Veronica Ghini Marzia Calvanese Andrea Colarusso Concetta Lauro Ermenegilda Parrilli Maria Luisa Tutino |
| author_facet | Marco Fondi Stefano Gonzi Mikolaj Dziurzynski Paola Turano Veronica Ghini Marzia Calvanese Andrea Colarusso Concetta Lauro Ermenegilda Parrilli Maria Luisa Tutino |
| author_sort | Marco Fondi |
| collection | DOAJ |
| description | hCDKL5 refers to the human cyclin-dependent kinase like 5 that is primarily expressed in the brain. Mutations in its coding sequence are often causative of hCDKL5 deficiency disorder, a devastating neurodevelopmental disorder currently lacking a cure. The large-scale recombinant production of hCDKL5 is desirable to boost the translation of preclinical therapeutic approaches into the clinic. However, this is hampered by the intrinsically disordered nature of almost two-thirds of the hCDKL5 sequence, making this region more susceptible to proteolytic attack, and the observed toxicity when the enzyme is accumulated in the cytoplasm of eukaryotic host cells. The bacterium <i>Pseudoalteromonas haloplanktis</i> TAC125 (PhTAC125) is the only prokaryotic host in which the full-length production of hCDKL5 has been demonstrated. To date, a system-level understanding of the metabolic burden imposed by hCDKL5 production is missing, although it would be crucial for upscaling of the production process. Here, we combined experimental data on protein production and nutrients assimilation with metabolic modelling to infer the global consequences of hCDKL5 production in PhTAC125 and to identify potential overproduction targets. Our analyses showed a remarkable accuracy of the model in simulating the recombinant strain phenotype and also identified priority targets for optimised protein production. |
| first_indexed | 2024-03-10T08:36:03Z |
| format | Article |
| id | doaj.art-4db893e695ef4f559a160a101cbcb545 |
| institution | Directory Open Access Journal |
| issn | 2218-1989 |
| language | English |
| last_indexed | 2024-03-10T08:36:03Z |
| publishDate | 2021-07-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Metabolites |
| spelling | doaj.art-4db893e695ef4f559a160a101cbcb5452023-11-22T08:39:03ZengMDPI AGMetabolites2218-19892021-07-0111849110.3390/metabo11080491Modelling hCDKL5 Heterologous Expression in BacteriaMarco Fondi0Stefano Gonzi1Mikolaj Dziurzynski2Paola Turano3Veronica Ghini4Marzia Calvanese5Andrea Colarusso6Concetta Lauro7Ermenegilda Parrilli8Maria Luisa Tutino9Department of Biology, University of Florence, Sesto F.no Florence, 50019 Florence, ItalyDepartment of Biology, University of Florence, Sesto F.no Florence, 50019 Florence, ItalyDepartment of Environmental Microbiology and Biotechnology, Institute of Microbiology, Faculty of Biology, University of Warsaw, 02-096 Warsaw, PolandMagnetic Resonance Center (CERM) and Department of Chemistry “Ugo Schiff”, University of Florence, via Sacconi 6, Sesto Fiorentino, 50019 Fiorentino, ItalyMagnetic Resonance Center (CERM) and Department of Chemistry “Ugo Schiff”, University of Florence, via Sacconi 6, Sesto Fiorentino, 50019 Fiorentino, ItalyDipartimento di Scienze Chimiche, Complesso Universitario Monte Sant’Angelo, 80126 Napoli, ItalyDipartimento di Scienze Chimiche, Complesso Universitario Monte Sant’Angelo, 80126 Napoli, ItalyDipartimento di Scienze Chimiche, Complesso Universitario Monte Sant’Angelo, 80126 Napoli, ItalyDipartimento di Scienze Chimiche, Complesso Universitario Monte Sant’Angelo, 80126 Napoli, ItalyDipartimento di Scienze Chimiche, Complesso Universitario Monte Sant’Angelo, 80126 Napoli, ItalyhCDKL5 refers to the human cyclin-dependent kinase like 5 that is primarily expressed in the brain. Mutations in its coding sequence are often causative of hCDKL5 deficiency disorder, a devastating neurodevelopmental disorder currently lacking a cure. The large-scale recombinant production of hCDKL5 is desirable to boost the translation of preclinical therapeutic approaches into the clinic. However, this is hampered by the intrinsically disordered nature of almost two-thirds of the hCDKL5 sequence, making this region more susceptible to proteolytic attack, and the observed toxicity when the enzyme is accumulated in the cytoplasm of eukaryotic host cells. The bacterium <i>Pseudoalteromonas haloplanktis</i> TAC125 (PhTAC125) is the only prokaryotic host in which the full-length production of hCDKL5 has been demonstrated. To date, a system-level understanding of the metabolic burden imposed by hCDKL5 production is missing, although it would be crucial for upscaling of the production process. Here, we combined experimental data on protein production and nutrients assimilation with metabolic modelling to infer the global consequences of hCDKL5 production in PhTAC125 and to identify potential overproduction targets. Our analyses showed a remarkable accuracy of the model in simulating the recombinant strain phenotype and also identified priority targets for optimised protein production.https://www.mdpi.com/2218-1989/11/8/491CDKL5genome-scale metabolic modellingprotein production |
| spellingShingle | Marco Fondi Stefano Gonzi Mikolaj Dziurzynski Paola Turano Veronica Ghini Marzia Calvanese Andrea Colarusso Concetta Lauro Ermenegilda Parrilli Maria Luisa Tutino Modelling hCDKL5 Heterologous Expression in Bacteria Metabolites CDKL5 genome-scale metabolic modelling protein production |
| title | Modelling hCDKL5 Heterologous Expression in Bacteria |
| title_full | Modelling hCDKL5 Heterologous Expression in Bacteria |
| title_fullStr | Modelling hCDKL5 Heterologous Expression in Bacteria |
| title_full_unstemmed | Modelling hCDKL5 Heterologous Expression in Bacteria |
| title_short | Modelling hCDKL5 Heterologous Expression in Bacteria |
| title_sort | modelling hcdkl5 heterologous expression in bacteria |
| topic | CDKL5 genome-scale metabolic modelling protein production |
| url | https://www.mdpi.com/2218-1989/11/8/491 |
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