About: Tat Peptide-Mediated Soluble Expression of the Membrane Protein LSECtin-CRD in Escherichia coli

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About: Tat Peptide-Mediated Soluble Expression of the Membrane Protein LSECtin-CRD in Escherichia coli Goto Sponge NotDistinct Permalink

An Entity of Type : schema:ScholarlyArticle, within Data Space : covidontheweb.inria.fr associated with source document(s)

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type	Academic Article research paper schema:ScholarlyArticle
isDefinedBy	Covid-on-the-Web dataset
has title	Tat Peptide-Mediated Soluble Expression of the Membrane Protein LSECtin-CRD in Escherichia coli
Creator	Cheng, Li Tang, Li Cong, Jianbo Dong, Guofu Wang, Changzhen Wang, Mingqun Wu, Ke Wu, Yonghong Zhang, Chenggang Zhao, Pengkai
Source	PMC
abstract	The human liver and lymph node sinusoidal endothelial cell C-type lectin (hLSECtin), a type II integral membrane protein, containing a Ca(2+)-dependent carbohydrate recognition domain (CRD), has a well-established biological activity, yet its three-dimensional structure is unknown due to low expression yields and aggregation into inclusion bodies. Previous study has demonstrated that the HIV-1 virus-encoded Tat peptide (‘YGRKKRRQRRR’) can increase the yields and the solubility of heterologous proteins. However, whether the Tat peptide could promote the high-yield and soluble expression of membrane proteins in Escherichia coli is not known. Therefore, the prokaryotic expression vector pET28b-Tat-hLSECtin-CRD (using pET28b and pET28b-hLSECtin-CRD as controls) was constructed, and transformed into E. coli BL21 (DE3) cells and induced with isopropyl-β-d-thiogalactoside (IPTG) followed with identifying by SDS-PAGE and Western blot. Subsequently, the bacterial subcellular structure, in which overexpressed the heterologous proteins Tat-hLSECtin-CRD and Tat-free hLSECtin-CRD, was analyzed by transmission electron microscope (TEM) respectively, and the mannose-binding activity of Tat-hLSECtin-CRD was also determined. Expectedly, the solubility of Tat-LSECtin-CRD significantly increased compared to Tat-free LSECtin-CRD (**p < 0.01) with prolonged time, and the Tat-LSECtin-CRD had a significant mannose-binding activity. The subcellular structure analysis indicated that the bacterial cells overexpressed Tat-hLSECtin-CRD exhibited denser region compared with controls, while dot denser region aggregated in the two ends of bacterial cells overexpressed Tat-free hLSECtin-CRD. This study provided a novel method for improving the soluble expression of membrane proteins in prokaryotic systems by fusion with the Tat peptide, which may be potentially expanded to the expression of other membrane proteins.
has issue date	2013-12-16(xsd:dateTime)
bibo:doi	10.1371/journal.pone.0083579
bibo:pmid	24358298
has license	cc-by
sha1sum (hex)	046a56d635edc933ed8f30ccf1ae9fd488dfe645
schema:url	https://doi.org/10.1371/journal.pone.0083579
resource representing a document's title	Tat Peptide-Mediated Soluble Expression of the Membrane Protein LSECtin-CRD in Escherichia coli
has PubMed Central identifier	PMC3865297
has PubMed identifier	24358298
schema:publication	PLoS One
resource representing a document's body	covid:046a56d635edc933ed8f30ccf1ae9fd488dfe645#body_text
is schema:about of	named entity 'However' named entity 'solubility' named entity 'subcellular' named entity 'soluble' named entity 'solubility' named entity 'DETERMINED' named entity 'DOT' named entity 'AGGREGATED' named entity 'SYSTEMS' named entity 'THIOGALACTOSIDE' named entity 'HETEROLOGOUS' named entity 'METHOD' named entity 'EXPRESSION' named entity 'PROLONGED' named entity 'MAY BE' named entity 'PROMOTE' named entity 'EXPRESSION VECTOR' named entity 'INCLUSION BODIES' named entity 'TAT PEPTIDE' named entity 'INCREASE' named entity 'UNKNOWN' named entity 'STUDY' named entity 'TAT' named entity 'TYPE II' named entity 'SOLUBLE' named entity 'RECOGNITION' named entity 'ESTABLISHED' named entity 'MEMBRANE PROTEINS' named entity 'INDUCED' named entity 'LECTIN' named entity 'SOLUBILITY' named entity 'YIELD' named entity 'AGGREGATION' named entity 'CRD' named entity 'PROVIDED' named entity 'NOT KNOWN' named entity 'SIGNIFICANT' named entity 'COMPARED' named entity 'DUE TO' named entity 'CONTAINING' named entity 'BIOLOGICAL ACTIVITY' named entity 'BL21' named entity 'CARBOHYDRATE' named entity 'HIV-1' named entity 'HUMAN LIVER' named entity 'PROKARYOTIC' named entity 'TAT PEPTIDE' named entity 'PROTEIN ' named entity 'HIGH' named entity 'INCREASED' named entity 'FOLLOWED' named entity 'TYPE' named entity 'TIME' named entity 'SDS-PAGE' named entity 'ANALYZED' named entity 'LOW EXPRESSION' named entity 'REGION' named entity 'DEMONSTRATED' named entity 'LYMPH NODE' named entity 'SINUSOIDAL' named entity 'IDENTIFYING' named entity 'ENDS' named entity 'SOLUBLE' named entity 'MEMBRANE PROTEIN' named entity 'MEDIATED' named entity 'LSECTIN' named entity 'ENDOTHELIAL CELL' named entity 'CELL C' named entity 'CONSTRUCTED' named entity 'DOMAIN' named entity 'WESTERN BLOT' named entity 'PROKARYOTIC EXPRESSION'

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