Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UNIPROT:P11021 (BiP)
 2,049 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Constructing eukaryotic expressing vector of pMT/Bip/V5-His A-HIV gp120 and transfecting S2 cells to establish stable gp120-expressing S2 cell line. The gp120 gene of HIV CRF07-BC epidemic in China was amplified by polymerase chain reaction from the recombinant vector PRSV-gp120 and confirmed by DNA sequence analysis. The DNA fragment of HIV gp120 was digested with the restriction endonucleases NcoI and XhoI, then inserted into eukaryotic expressing vector pMT/BiP/V5-His A. A selection vector containing the streptomyces griseochromogenes bsd gene conferring blasticidin resistance was cotransfected into drosophila S2 cells by Cellfectin II reagent. The stable cell line was established following repeated screening by blasticidin. HIV gp120 was purified by a Ni-NTA affinity column followed by molecular sieve chromatography. Recombinant protein was characterized by SDS-PAGE, Western blot and ELISA. The eukaryotic expressing vector pMT/BiP/V5-His A-HIV gp120 was constructed, stable expressing cell line was established, and protein was expressed and purified successfully. This result will contribute to functional study of gp120 and vaccine design against AIDS.
 ...
PMID:[Establishment of stable cell line and expression and purification of HIV gp120 in Drosophila S2 cells]. 2134 50

Plants show great potential for producing recombinant proteins in a cost-effective manner. Many strategies have therefore been employed to express high levels of recombinant proteins in plants. Although foreign domains are fused to target proteins for high expression or as an affinity tag for purification, the retention of foreign domains on a target protein may be undesirable, especially for biomedical purposes. Thus, their removal is often crucial at a certain time point after translation. Here, we developed a new strategy to produce target proteins without foreign domains. This involved in vivo removal of foreign domains fused to the N-terminus by the small ubiquitin-related modifier (SUMO) domain/SUMO-specific protease system. This strategy was tested successfully by generating a recombinant gene, BiP:p38:bdSUMO : His:hLIF, that produced human leukemia inhibitory factor (hLIF) fused to p38, a coat protein of the Turnip crinkle virus; the inclusion of p38 increased levels of protein expression. The recombinant protein was expressed at high levels in the leaf tissue of Nicotiana benthamiana. Coexpression of bdSENP1, a SUMO-specific protease, proteolytically released His:hLIF from the full-length recombinant protein in the endoplasmic reticulum of N. benthamiana leaf cells. His:hLIF was purified from leaf extracts via Ni2+-NTA affinity purification resulting in a yield of 32.49 mg/kg, and the N-terminal 5-residues were verified by amino acid sequencing. Plant-produced His:hLIF was able to maintain the pluripotency of mouse embryonic stem cells. This technique thus provides a novel method of removing foreign domains from a target protein in planta.
 ...
PMID:In Vivo Removal of N-Terminal Fusion Domains From Recombinant Target Proteins Produced in Nicotiana benthamiana. 3232 82