For better understanding the chemical or biological information of ZNF191 （243-368）, we expressed the fusion protein of GST and ZNF191 （243-368）, and used it to obtain the binding DNA sequence of this zinc finger ...For better understanding the chemical or biological information of ZNF191 （243-368）, we expressed the fusion protein of GST and ZNF191 （243-368）, and used it to obtain the binding DNA sequence of this zinc finger protein. But in the process of expression and purification, we found this fusion protein slowly degradated. For resolving this problem, we simultaneously added charged amino acids L-Arg and L-Glu to the solution of fusion protein, and demonstrated that this method can dramatically increase the stability of this fusion protein. This method can make the fusion protein suitable for the continuous works, especially for situations where high protein concentration and long-term stability without precipitate and degradation of protein are required.展开更多
Two mutants of the zinc finger protein, ZNF191(243—368) I323W and R327W, are suc-cessfully obtained by site-directed mutagenesis. The fluorescence spectrum is used to study the interaction between these two mutants a...Two mutants of the zinc finger protein, ZNF191(243—368) I323W and R327W, are suc-cessfully obtained by site-directed mutagenesis. The fluorescence spectrum is used to study the interaction between these two mutants and the oligonucleotides. The influence of the mutation on the interaction has been studied using ethidium bromide (EB) as the fluorescence probe. The binding constants of the I323W-DNA and R327W-DNA have been calculated and the possible binding models have been discussed.展开更多
AIM:Human zinc finger protein 191 (ZNF191) was cloned and characterized as a Krueppel-like transcription factor, which might be relevant to many diseases such as liver cancer,neuropsychiatric and cardiovascular diseas...AIM:Human zinc finger protein 191 (ZNF191) was cloned and characterized as a Krueppel-like transcription factor, which might be relevant to many diseases such as liver cancer,neuropsychiatric and cardiovascular diseases. Although progress has been made recently, the biological function of ZNF191 remains largely unidentified. The aim of this study was to establish a ZNF191 transgenic mouse model,which would promote the functional study of ZNF191.METHODS:Transgene fragments were microinjected into fertilized eggs of mice.The manipulated embryos were transferred into the oviducts of pseudo-pregnant female mice.The offsprings were identified by PCR and Southern blot analysis.ZNF191 gene expression was analyzed by RT-PCR.Transgenic founder mice were used to establish transgenic mouse lineages.The first generation (F1) and the second generation (F2) mice were identified by PCR analysis.Ten-week transgenic mice were used for pathological examination.RESULTS: Four mice were identified as carrying copies of ZNF191 gene.The results of RT-PCR showed that ZNF191 gene was expressed in the liver,testis and brain in one of the transgenic mouse lineages.Genetic analysis of transgenic mice demonstrated that ZNF191 gene was integrated into the chromosome at a single site and could be transmitted stably. Pathological analysis showed that the expression of ZNF 191 did not cause obvious pathological changes in multiple tissues of transgenic mice.CONCLUSION:ZNF191 transgenic mouse model would facilitate the investigation of biological functions of ZNF191 in vivo.展开更多
文摘For better understanding the chemical or biological information of ZNF191 （243-368）, we expressed the fusion protein of GST and ZNF191 （243-368）, and used it to obtain the binding DNA sequence of this zinc finger protein. But in the process of expression and purification, we found this fusion protein slowly degradated. For resolving this problem, we simultaneously added charged amino acids L-Arg and L-Glu to the solution of fusion protein, and demonstrated that this method can dramatically increase the stability of this fusion protein. This method can make the fusion protein suitable for the continuous works, especially for situations where high protein concentration and long-term stability without precipitate and degradation of protein are required.
基金This work was supported by the National Natural Science Foundation of China (Grant Nos. 30170228 and 39990600).
文摘Two mutants of the zinc finger protein, ZNF191(243—368) I323W and R327W, are suc-cessfully obtained by site-directed mutagenesis. The fluorescence spectrum is used to study the interaction between these two mutants and the oligonucleotides. The influence of the mutation on the interaction has been studied using ethidium bromide (EB) as the fluorescence probe. The binding constants of the I323W-DNA and R327W-DNA have been calculated and the possible binding models have been discussed.
基金Supported by the National Natural Science Foundation of China,No.39830360
文摘AIM:Human zinc finger protein 191 (ZNF191) was cloned and characterized as a Krueppel-like transcription factor, which might be relevant to many diseases such as liver cancer,neuropsychiatric and cardiovascular diseases. Although progress has been made recently, the biological function of ZNF191 remains largely unidentified. The aim of this study was to establish a ZNF191 transgenic mouse model,which would promote the functional study of ZNF191.METHODS:Transgene fragments were microinjected into fertilized eggs of mice.The manipulated embryos were transferred into the oviducts of pseudo-pregnant female mice.The offsprings were identified by PCR and Southern blot analysis.ZNF191 gene expression was analyzed by RT-PCR.Transgenic founder mice were used to establish transgenic mouse lineages.The first generation (F1) and the second generation (F2) mice were identified by PCR analysis.Ten-week transgenic mice were used for pathological examination.RESULTS: Four mice were identified as carrying copies of ZNF191 gene.The results of RT-PCR showed that ZNF191 gene was expressed in the liver,testis and brain in one of the transgenic mouse lineages.Genetic analysis of transgenic mice demonstrated that ZNF191 gene was integrated into the chromosome at a single site and could be transmitted stably. Pathological analysis showed that the expression of ZNF 191 did not cause obvious pathological changes in multiple tissues of transgenic mice.CONCLUSION:ZNF191 transgenic mouse model would facilitate the investigation of biological functions of ZNF191 in vivo.
