pFUSE-hlgG1e1-FC2 BioVector NTCC质粒载体菌种细胞基因保藏中心

pFUSE-hlgG1e1-FC2 BioVector NTCC质粒载体菌种细胞基因保藏中心 GENERAL PRODUCT USE
pFUSE-Fc is a family of plasmid developed to facilitate the construction
of Fc-fusion proteins by fusing the effector region of a protein to the Fc
region of an immunoglobul in G (IgG).
pFUSE-Fc plasmids yield high levels of Fc-fusion proteins. The level of
expression is usually in the µg/mL range. They can be transfected in a
variety of mammalian cells, including myeloma cell lines, CHO cells,
monkey COS cells and human embryonic kidney (HEK)293 cells, cells
that are commonly used in protein purification systems.
pFUSE-Fc2 (IL2ss) plasmids allow the secretion of Fc-Fusion proteins.
They contain the IL2 signal sequence (IL2ss) for the generation of FcFusion
proteins derived from proteins that are not naturally secreted. As FcFusion
proteins are secreted, they can be easily detected in the supernatant
of pFUSE-Fc-transfected cells by SDS-PAGE. Furthermore, functional
domains can be identified by immunoblotting and ligand blotting.
Fc-Fusion proteins can be easily purified by single-step protein A o r
protein G affinity chromatography.
InvivoGen provides pFUSE-Fc vectors featuring Fc regions from
d i fferent species and isotypes. In humans, there are four isotypes: Ig G 1 ,
IgG2, IgG3 and IgG4. The Fc region mediates effector functions, such
as antibody-dependent cellular cytotoxicity (ADCC) and complementdependent
cytotoxicity (CDC). IgG isoforms exert different levels of
e ffector functions increasing in the order of IgG4Human IgG1 dispays high ADCC and CDC, and is the most suitable for
therapeutic use against pathogens and cancer cells.
Under certain circumstances, for example when depletion of the targ e t
cell is undesirable, abrogating effector functions is required. On the
c o n t r a r y, in the case of antibodies intended for oncology use, increasing
e ffector functions may improve their therapeutic activity1
. Modifying
e ffector functions can be achieved by engineering the Fc regions to
either improve or reduce their binding to FcγRs or the complement
factors. Amino acids substitutions have been made in the human IgG1
Fc region in order to increase or reduce its ADCC and CDC.
PLASMID FEATURES
• hIgG1e1-Fc (human IgG1 engineered Fc): The Fc region comprises the
CH2 and CH3 domains of the IgG heavy chain and the hinge region. T h e
hinge serves as a flexible spacer between the two parts of the Fc-fusion
protein, allowing each part of the molecule to function independently.
The Fc region binds to neonatal FcR (FcRn), a receptor expressed on the
surface of endothelial cells. This interaction, which is pH-dependent,
protects the IgG from lysosomal degradation thus mediating the serum
persistence of IgG antibodies. The human IgG1 Fc domain was engineered
by introducing mutations in the FcRn binding sites leading to higher FcRn
binding affinity at pH 6.02
. The engineered hIgG1e2 Fc contains two amino
acid substitutions: T250Q and M428L.
• hEF1-HTLV prom is a composite promoter comprising the Elongation
Factor-1α (EF-1α) core promoter3 and the R segment and part of the U5
sequence (R-U5’) of the Human T-Cell Leukemia Virus (HTLV) Type 1
Long Terminal Repeat4
. The EF-1α promoter exhibits a strong activity
and yields long lasting expression of a transgene in vivo. The R-U5’ has
been coupled to the EF-1α core promoter to enhance stability of RNA.
• IL2 ss: The IL2 signal sequence contains 20 amino acids and share
common characteristics with signal peptides of other secretory proteins. T h e
intracellular cleavage of the IL2 signal peptide occurs after Ser20 and leads to
the secretion of the antigenic protein.
• MCS: The multiple cloning site contains several restriction sites that are
compatible with many other enzymes, thus facilitating cloning.
• SV40 pAn: the Simian Virus 40 late polyadenylation signal enables
efficient cleavage and polyadenylation reactions resulting in high levels
of steady-state mRNA5
.
• ori: a minimal E. coli origin of replication to limit vector size, but with
the same activity as the longer Ori.
• CMV enh / hFerL prom: This composite promoter combines the
human cytomegalovirus immediate-early gene 1 enhancer and the core
promoter of the human ferritin light chain gene. This ubiquitous promoter
drives the expression of the Zeocin™-resistance gene in mammalian cells.
• EM2KC is a bacterial promoter that enables the constitutive expression
of the antibiotic resistance gene in E. coli. EM2KC is located within an
intron and is spliced out in mammalian cells.
• Zeo: Resistance to Zeocin™ is conferred by the Sh ble gene from
S t reptoalloteichus hindustanus The same resistance gene confers
selection in both mammalian cells and E. coli.
• ßGlo pAn: The human beta-globin 3’UTR and polyadenylation
sequence allows efficient arrest of the transgene transcription6
.BioVector NTCC质粒载体菌种细胞蛋白抗体基因保藏中心www.biovector.net [Supplier来源] http://www.biovector.net