Browse
Submit Data
Databases
API
Help

Dataset Information

9 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Enhanced Recombinant Protein Production of Soluble, Highly Active and Immobilizable PNGase F.

ABSTRACT: High resolution analysis of N-glycans can be performed after their endoglycosidase mediated removal from proteins. N-glycosidase F peptide (PNGase F) is one the most frequently used enzyme for this purpose. Because of the significant demand for PNGase F both in basic and applied research, rapid and inexpensive methods are of great demand for its large-scale production, preferably in immobilizable form to solid supports or surfaces. In this paper, we report on the high-yield production of N-terminal 6His-PNGase F enzyme in a bacterial Escherichia coli SHuffle expression system. The activity profile of the generated enzyme was compared to commercially available PNGase F enzymes, featuring higher activity for the former. The method described here is thus suitable for the cost-effective production of PNGase F in an active, immobilizable form.

SUBMITTER: Kovacs N

PROVIDER: S-EPMC9259526 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Discovery of Highly Active Recombinant PNGase H+ Variants Through the Rational Exploration of Unstudied Acidobacterial Genomes.

Project description: Not available

| S-EPMC7348039 | biostudies-literature

Production of Highly Active Recombinant Dermonecrotic Toxin of Bordetella Pertussis.

Project description: Not available

| S-EPMC7551409 | biostudies-literature

Fusion Tag Design Influences Soluble Recombinant Protein Production in <i>Escherichia coli</i>.

Project description: Not available

| S-EPMC9321918 | biostudies-literature

Novel Highly Soluble Chimeric Recombinant Spidroins with High Yield.

Project description: Not available

| S-EPMC7554824 | biostudies-literature

Escherichia coli Can Adapt Its Protein Translocation Machinery for Enhanced Periplasmic Recombinant Protein Production.

Project description: Not available

| S-EPMC7000420 | biostudies-literature

A Rational Approach for the Production of Highly Soluble and Functional Sunflower Protein Hydrolysates.

Project description: Not available

| S-EPMC8003716 | biostudies-literature

Omics Analysis Reveals the Mechanism of Enhanced Recombinant Protein Production Under Simulated Microgravity.

Project description: Not available

| S-EPMC7044180 | biostudies-literature

Escherichia coli can adapt its protein translocation machinery for enhanced periplasmic recombinant protein production

Project description:Recently, we engineered a tunable rhamnose promoter-based setup for the production of recombinant proteins in E. coli. This setup enabled us to show that being able to precisely set the production rate of a secretory recombinant protein is critical to enhance protein production yields in the periplasm. It is assumed that precisely setting the production rate of a secretory recombinant protein is required to harmonize its production rate with the protein translocation capacity of the cell. Here, using proteome analysis we show that enhancing periplasmic production of human Growth Hormone (hGH) using the tunable rhamnose promoter-based setup is accompanied by increased accumulation levels of at least three key players in protein translocation; the peripheral motor of the Sec-translocon SecA, leader peptidase (LepB) and the cytoplasmic membrane protein integrase/chaperone YidC. Thus, enhancing periplasmic hGH production leads to increased Sec-translocon capacity, increased capacity to cleave signal peptides from secretory proteins and an increased capacity of an alternative membrane protein biogenesis pathway, which frees up Sec-translocon capacity for protein secretion. When cells with enhanced periplasmic hGH production yields were harvested and subsequently cultured in the absence of inducer, SecA, LepB and YidC levels went down again. This indicates that when using the tunable rhamnose-promoter system to enhance the production of a protein in the periplasm, E. coli can adapt its protein translocation machinery for enhanced recombinant protein production in the periplasm.

2020-03-05 | PXD013168 | Pride

Multiplex secretome engineering enhances recombinant protein production and purity

Project description:Host cell proteins (HCPs) are process-related impurities generated during biotherapeutic protein production. HCPs can be problematic if they pose a significant metabolic demand, degrade product quality, or contaminate the final product. Here, we present an effort to create a “clean” Chinese hamster ovary (CHO) cell by disrupting multiple genes to eliminate HCPs. Using a model of CHO cell protein secretion, we predicted the elimination of unnecessary HCPs could have a non-negligible impact on protein production. We analyzed the total HCP content of 6-protein, 11-protein, and 14-protein knockout clones and characterized their growth in shake flasks and bioreactors. These cell lines exhibited a substantial reduction in total HCP content (40%-70%). We also observed higher productivity and improved growth characteristics, in specific clones. With the reduced HCP content, protein A and ion exchange chromatography more efficiently purified a monoclonal antibody (mAb) produced in these cells during a three-step purification process. Thus, substantial improvements can be made in protein titer and purity through large-scale HCP deletion, providing an avenue to increased quality and affordability of high-value biopharmaceuticals.

2020-03-19 | GSE144624 | GEO

Hyper-enhanced production of foreign recombinant protein by fusion with the partial polyhedrin of nucleopolyhedrovirus.

Project description: Not available

| S-EPMC3621880 | biostudies-literature