Interdisciplinary gene manipulation, molecular cloning, and recombinant expression of modified human growth hormone isoform-1 in E. coli system.

Batch fermentation Escherichia coli Growth hormone Immunodetection MALDI-TOF Nano-LC-ESI MS/MS Proliferation activity Recombinant drug Vero cells

Journal

International journal of biological macromolecules

ISSN: 1879-0003

Titre abrégé: Int J Biol Macromol

Pays: Netherlands

ID NLM: 7909578

Informations de publication

Date de publication:
06 Dec 2023

Historique:

received: 06 09 2023

revised: 25 11 2023

accepted: 03 12 2023

pubmed: 8 12 2023

medline: 8 12 2023

entrez: 7 12 2023

Statut: aheadofprint

Résumé

Growth hormone (GH) is a hormone that promotes growth, cell reproduction, and cell restoration in humans and animals. Production of recombinant human growth hormone (rhGH) in Escherichia coli (E. coli) and assessment of its characteristics and proliferation stimulatory activity. The hGH gene was cloned into a pET 3a expression vector and transformed into a competent E. coli cell. The refolded hGH was purified, Western blot and batch fermentation were performed. Cell cytotoxicity was tested on Vero cells, and MALDI-TOF and Nano-LC-ESI MS/MS were used for protein and target peptide analysis. Induced rhGH was purified with a concentration of 511.9 mg/ml. Western blot confirmed the molecular identity of rhGH, showing a single 22 kDa band. The bacterial growth at OD Biologically active native rhGH protein was successfully expressed in the Prokaryotic system. Our goal is to increase its production on a pilot level in the native form at a high activity effect identical to isoform 1.

Sections du résumé

BACKGROUND BACKGROUND

Growth hormone (GH) is a hormone that promotes growth, cell reproduction, and cell restoration in humans and animals.

OBJECTIVES OBJECTIVE

Production of recombinant human growth hormone (rhGH) in Escherichia coli (E. coli) and assessment of its characteristics and proliferation stimulatory activity.

METHODS METHODS

The hGH gene was cloned into a pET 3a expression vector and transformed into a competent E. coli cell. The refolded hGH was purified, Western blot and batch fermentation were performed. Cell cytotoxicity was tested on Vero cells, and MALDI-TOF and Nano-LC-ESI MS/MS were used for protein and target peptide analysis.

RESULTS RESULTS

Induced rhGH was purified with a concentration of 511.9 mg/ml. Western blot confirmed the molecular identity of rhGH, showing a single 22 kDa band. The bacterial growth at OD

CONCLUSION CONCLUSIONS

Biologically active native rhGH protein was successfully expressed in the Prokaryotic system. Our goal is to increase its production on a pilot level in the native form at a high activity effect identical to isoform 1.

Identifiants

DOI: 10.1016/j.ijbiomac.2023.128637 PMID: 38061513

pubmed: 38061513

pii: S0141-8130(23)05536-8

doi: 10.1016/j.ijbiomac.2023.128637

pii:

doi:

Types de publication

Journal Article

Langues

eng

Sous-ensembles de citation

Pagination

128637

Informations de copyright

Déclaration de conflit d'intérêts

Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Dr. Sami Mohamed Nasr reports financial support, administrative support, equipment, drugs, or supplies, and travel were provided by Science and Technology Development Fund. Dr. Sami Mohamed Nasr reports a relationship with Theodor Bilharz Research Institute that includes: employment. Co-author Dr. Hend Okasha and member Dr. Safia Samir are employee at Theodor Bilharz research institute.