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Protocol for producing brain-derived neurotrophic factor and neurotrophin-4 in their pro and active form in Escherichia coli

Year: 2025
Type: article
Abstract: Brain-derived neurotrophic factor (BDNF) and neurotrophin-4 (NT4) are neurotrophic growth factors, signaling primarily via the tropomyosin receptor kinase B (TrkB). Here, we present a protocol for producing BDNF and NT4 in their pro and active form in Escherichia coli . We describe steps for E. coli expression, oxidative in vitro folding from inclusion bodies, and purification. We then detail procedures for proteolytic activation and characterization of the purified neurotrophins. For complete details on the use and execution of this protocol, please refer to Holzner et al. 1 • Instructions for producing proBDNF and proNT4 inclusion bodies in E. coli • Steps for in vitro folding of proBDNF and proNT4 with intact disulfide bridges • Guidance on batch purification and activation of folded proBDNF and proNT4 Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Brain-derived neurotrophic factor (BDNF) and neurotrophin-4 (NT4) are neurotrophic growth factors, signaling primarily via the tropomyosin receptor kinase B (TrkB). Here, we present a protocol for producing BDNF and NT4 in their pro and active form in Escherichia coli . We describe steps for E. coli expression, oxidative in vitro folding from inclusion bodies, and purification. We then detail procedures for proteolytic activation and characterization of the purified neurotrophins.
DOI: 10.1016/j.xpro.2025.103715
Language: English
Open Access status: gold
Estimated APC paid: $1,550
OpenAlex ID: W4409018754
FWCI:
Cites: 4
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Related to: 10