PUBLICATION
Recovery of recombinant zebrafish p53 protein from inclusion bodies and its binding activity to p53 mRNA in vitro
- Authors
- Zhao, X., Liu, M., Wu, N., Ding, L., Liu, H., and Lin, X.
- ID
- ZDB-PUB-100408-22
- Date
- 2010
- Source
- Protein Expression and Purification 72(2): 262-266 (Journal)
- Registered Authors
- Keywords
- p53, Zebrafish, RNA binding protein, Protein expression, Inclusion body
- MeSH Terms
-
- Animals
- Electrophoresis, Polyacrylamide Gel
- Embryo, Nonmammalian/metabolism
- Escherichia coli/genetics
- Escherichia coli/metabolism
- Inclusion Bodies/chemistry*
- RNA, Messenger/genetics
- RNA, Messenger/metabolism*
- Recombinant Proteins/genetics
- Recombinant Proteins/isolation & purification
- Recombinant Proteins/metabolism
- Tumor Suppressor Protein p53/genetics
- Tumor Suppressor Protein p53/isolation & purification*
- Tumor Suppressor Protein p53/metabolism*
- Zebrafish/genetics
- Zebrafish Proteins/genetics
- Zebrafish Proteins/isolation & purification*
- Zebrafish Proteins/metabolism*
- PubMed
- 20363330 Full text @ Protein Expr. Purif.
Citation
Zhao, X., Liu, M., Wu, N., Ding, L., Liu, H., and Lin, X. (2010) Recovery of recombinant zebrafish p53 protein from inclusion bodies and its binding activity to p53 mRNA in vitro. Protein Expression and Purification. 72(2):262-266.
Abstract
p53 protein is an important regulation factor that can bind to p53 mRNA to regulate its translation in human and murine. To determine if a similar interaction exists in zebrafish and if the interaction affects zebrafish development, we cloned and expressed p53 protein from zebrafish in Escherichia coli. Soluble p53 protein with high purity was successfully obtained using the optimized renaturation approach. Results of a UV-crosslinking experiment and immunoprecipitation:RT-PCR analysis confirmed that the purified p53 protein could bind specifically to its cognate mRNA. Our results suggest that selecting a suitable buffer is important for renaturing p53 protein from inclusion bodies. We also demonstrated a specific interaction between p53 and it own RNA in zebrafish. Measurement of the binding activity may be a useful approach for identifying the activity of recombinant p53 protein in vitro.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping