A plasmid system for high-level expression and in vitro processing of recombinant proteins

Gene ◽  
1993 ◽  
Vol 130 (1) ◽  
pp. 121-126 ◽  
Author(s):  
Johannes Pohlner ◽  
Joachim Krämer ◽  
Thomas F. Meyer
Keyword(s):  
Recombinant Proteins ◽  
High Level Expression ◽  
In Vitro Processing ◽  
High Level

scholarly journals High-Level Expression of Palmitoylated MPP1 Recombinant Protein in Mammalian Cells

Membranes ◽  
2021 ◽  
Vol 11 (9) ◽  
pp. 715
Author(s):  
Agnieszka Chytła ◽  
Weronika Gajdzik-Nowak ◽  
Agnieszka Biernatowska ◽  
Aleksander F. Sikorski ◽  
Aleksander Czogalla
Keyword(s):  
Recombinant Proteins ◽  
Mammalian Cells ◽  
High Yield ◽  
High Level Expression ◽  
Lateral Membrane ◽  
Protein Membrane ◽  
High Level ◽  
Lateral Organization

Our recent studies have pointed to an important role of the MAGUK family member, MPP1, as a crucial molecule interacting with flotillins and involved in the lateral organization of the erythroid plasma membrane. The palmitoylation of MPP1 seems to be an important element in this process; however, studies on the direct effect of palmitoylation on protein–protein or protein–membrane interactions in vitro are still challenging due to the difficulties in obtaining functional post-translationally modified recombinant proteins and the lack of comprehensive protocols for the purification of palmitoylated proteins. In this work, we present an optimized approach for the high-yield overexpression and purification of palmitoylated recombinant MPP1 protein in mammalian HEK-293F cells. The presented approach facilitates further studies on the molecular mechanism of lateral membrane organization and the functional impact of the palmitoylation of MPP1, which could also be carried out for other palmitoylated proteins.

scholarly journals High-level expression of IκB-β in the surface epithelium of the colon: in vitro evidence for an immunomodulatory role

1999 ◽  
Vol 66 (6) ◽  
pp. 1049-1056 ◽  
Author(s):  
Gary D. Wu ◽  
Ning Huang ◽  
Xiaoming Wen ◽  
Sue A. Keilbaugh ◽  
Hongyun Yang
Keyword(s):  
Surface Epithelium ◽  
High Level Expression ◽  
High Level

scholarly journals Thyroid hormone induces constitutive keratin gene expression during Xenopus laevis development.

1989 ◽  
Vol 9 (5) ◽  
pp. 1823-1831 ◽  
Author(s):  
P M Mathisen ◽  
L Miller
Keyword(s):  
Thyroid Hormone ◽  
Xenopus Laevis ◽  
Constitutive Expression ◽  
High Level Expression ◽  
Keratin Gene ◽  
Explant Cultures ◽  
Extensive Cell ◽  
High Level

We have used in vitro explant cultures of Xenopus laevis skin to investigate the role that the thyroid hormone triiodothyronine (T3) plays in activating the 63-kilodalton (kDa) keratin genes. The activation of these genes in vivo requires two distinct steps, one independent of T3 and one dependent on T3. In this report we have shown that the same two steps are required to fully activate the 63-kDa keratin genes in skin explant cultures, and we have characterized the T3-mediated step in greater detail. Unlike the induction of transcription by T3 or steroid hormones in adult tissues, there was a long latent period of approximately 2 days between the addition of T3 to skin cultures and an increase in concentration of keratin mRNA. While the T3 induction of 63-kDa keratin gene transcription cannot occur until age 48, a short transient exposure of stage 40 skin cultures to T3 resulted in high-level expression of these genes 5 days later, when normal siblings had reached stage 48. This result indicates that T3 induces a stable change in epidermal cells which can be expressed much later, after extensive cell proliferation has occurred in the absence of T3. Once the 63-kDa keratin genes were induced, they were stably expressed, and by the end of metamorphosis T3 had no further effect on their expression. The results suggest that T3 induces constitutive expression of the 63-kDa keratin genes during metamorphosis.

High-level expression of lipase in Escherichia coli and recovery of active recombinant enzyme through in vitro refolding

2010 ◽  
Vol 70 (1) ◽  
pp. 75-80 ◽  
Author(s):  
Neda Akbari ◽  
Khosro Khajeh ◽  
Sasan Rezaie ◽  
Saeed Mirdamadi ◽  
Mahmoud Shavandi ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Recombinant Enzyme ◽  
High Level Expression ◽  
High Level ◽  
In Vitro Refolding

scholarly journals Effects of Overexpressed P2X7 on Leukemia Progression in Mouse Acute Myeloid Leukemia Model

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 5141-5141
Author(s):  
Wenli Feng ◽  
Xiao Yang ◽  
Rong Wang ◽  
Feifei Yang ◽  
Hao Wang ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Mouse Model ◽  
Myeloid Leukemia ◽  
Leukemia Cells ◽  
High Level Expression ◽  
Innovation Fund ◽  
High Level ◽  
Acute Myeloid ◽  
Proliferation Potential

Abstract P2X7 is one of the P2X family ligand-gated ion channel receptors. High level expression of P2X7 is reported in various malignant cells. High level expression of P2X7 was also detected in leukemia patients, especially in relapsed cases. A hyposensitive P2X7 mutant, N187D P2X7, was cloned from leukemia cells. However, the role of P2X7 on the progression of leukemia was poorly understood. In the present study, we studied the effects of P2X7 receptor in acute myeloid leukemia mouse model. We established MLL-AF9 induced AML mouse model with high level expression of P2X7 (MA9-P2X7). High level of P2X7 significantly accelerated the progression of leukemia. Homing capability experiment demonstrated that MA9-P2X7 cells had lower homing potential. The apoptotic rate of freshly isolated leukemia cells had no significant difference between two groups. However, both in vitro culture experiments and in vivo BrdU incorporation assay demonstrated that MA9-P2X7 cells had enhanced proliferation potential, i.e. more S and G2/M phase cells but less G0/G1 phase cells. Moreover, upon treatment with Ara-C, though MA9-P2X7 cells were more sensitive to Ara-C, but the mice had shorter survival time. Furthermore, in vitro colony assay and limiting dilution transplantation experiments showed that MA9-P2X7 cells formed more colonies and had a 7.25-folds increase in LICs frequency. We also detected the expression of c-Kit, and the results showed that the majority of MA9-P2X7 cells were c-Kit+, whereas control cells have two populations and more than half of them were c-Kit-. Leukemia mice in MA9-P2X7 group had shorter survival time than those in the c-Kit+ control group. Our results suggested that leukemia cells overexpressing P2X7 possessed the characteristics of both greater proliferation potential and higher LICs frequency, which contributed to the accelerated progression of leukemia. This work was supported by grants 81570153, 81770183 and 81170511 from the National Natural Science Foundation of China (NSFC); programs 2016-I2M-2-006 and 2017-I2M-1-015 from the CAMS Innovation Fund for Medical Sciences (CIFMS); grant 17JCZDJC35000 from the Tianjin Natural Science Foundation; and Graduate Student Innovation Fund (2014-0710-1021) from Peking Union Medical College. Z.GG. is a recipient of the New Century Excellent Talents in University (NCET-08-0329). Disclosures No relevant conflicts of interest to declare.

scholarly journals High-level expression of CXCL1/GROα in uterine cervix cancer is linked to advanced stage and facilitates tumor cell malignant processes in an autocrine/paracrine manner

2020 ◽  
Author(s):  
XiaXia Man ◽  
XiaoLin Yang ◽  
ZhenTong Wei ◽  
YuYing Tan ◽  
WanYing Li ◽  
...  
Keyword(s):  
Hela Cells ◽  
Uterine Cervix ◽  
Human Cancer ◽  
Functional Relation ◽  
Cervix Cancer ◽  
High Level Expression ◽  
Uterine Cervix Cancer ◽  
And Migration ◽  
High Level

Abstract Background It has previously accepted that several types of human cancer constitutively express CXCL1, which may implicated in various aspects of tumors. Here, we sought to assess the expression and of CXCL1 in human uterine cervix cancer (UCC) and its potential role and mechanism in UCC. Methods CXCL1 protein expression in a uterine cervical tissue microarray was assessed by immunohistochemistry. The roles of CXCL1 on HeLa UCC cells were detected by CCK-8, transwell and apoptosis assays. Western blotting and ERK1/2 blocker PD98059 were utilized to explore whether ERK signal was implicated in CXCL1-mediated UCC processes. Results CXCL1 was expressed by HeLa, PHM1-41 cells as well as cervical tissues, in which UCC tissues showed an obviously high level of CXCL1 compared with non-cancerous tissues. Additionally, of the cancer tissues, CXCL1 high-level expression was notably relevant to poor clinical stages. In vitro, the growth and migration of HeLa cells were enhanced in the presence of exogenous CXCL1. Gain-function assays revealed that CXCL1 overexpression promoted growth and migration response to HeLa cells via autocrine/paracrine manners. Moreover, CXCL1 also led to the inhibition of apoptosis in HeLa cells. Finally, CXCL1 overexpression in HeLa cells influenced the expression of ERK signal-related genes including ERK, p-ERK, cyclin D1 and Bax, and cell malignant behaviors derived from CXCL1 overexpresion were further interrupt in the presence of PD98059. Conclusion Our findings provided the potential roles of CXCL1 as a promoter and the novel understanding of the functional relation of CXCL1 with ERK signal pathway in UCC.

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