Protein Expression

Each protein expression and purification project is unique. High levels of protein expression in the host of choice are essential for economical purification. We offer complete protein production services including gene synthesis, cloning and expression and protein purification.

The first critical step in our protein expression process is to design the optimal expression plasmid taking into consideration the structure and function of the expressed protein, host species, purification strategy, and purpose of protein production, incorporating tags and sequence elements as appropriate.  We determine the most appropriate expression vector for E. coli, yeast and mammalian cells, facilitating soluble protein expression and minimizing the number of purification steps.

We have extensive experience with a variety of components to design optimal protein expression plasmids:

> Episomal or Integrated with low or high copy number
> Inducible Promoters (T7, ara, lac, GAL, Tet-On)
> Constitutive Promoters (ADH, TEF, CMV, EF1, Ubiquitin)
> Purification Tags (HIS, FLAG, GST, STREP, MBP, CBP, CYD)
> Protease Sites (TEV, 3C, Thrombin, EK, Xa)
> Solubility Tags (TrxA, NusA, Z-domain, GB1, GST, MBP)
> Secretion Tags (DsbA, DsbC, PelB, IgG, Amylase)
> Epitope Tags (c-MYC, HA, HSV, FLAG, V5, VSV-G)
> Labeling Tags (ACP, BAP, GFP, Intein, SNAP, CLIP, Lumio)
> Co-expression (GroEL, DnaK, GrpE, Growth Factors)

Protein expression levels are also enhanced by optimizing:
> Growth conditions, temperature, and length of induction
> IPTG, arabinose, heat, cold, or auto-induction
> E. coli strains for disulfide bonds and toxic genes
>Transient or stable in CHO, HEK293, NSO, & COS
> Fermentation conditions for yeast

Protein Purification

One World Biotech is uniquely qualified to develop and deliver complex, multi-step protein purification methods that yield highly pure and biologically active proteins. We use customer-supplied methods, published protocols, or we will develop or optimize procedures in-house. We use a GE AKTA FPLC for purification and supply all electronic data reports.

> His-Tag (IMAC) and other Tag Purification
> Affinity Chromatography
> Ion Exchange Chromatography (IEC)
> Size Exclusion Chromatography (SEC, Gel Filtration)
> Hydrophobic Interaction Chromatography (HIC)
> Isoelectric Focusing (IEF)
> Preparative PAGE
> Inclusion Bodies Purification
> Renaturation of Denatured Proteins (Protein Folding)
> Endotoxin and Purification Tag Removal
> New Method Development

We offer the following tools for identification and analytical characterization of recombinant proteins:

> Protein assay and UV-Vis absorption spectrum
> SDS-PAGE, Native PAGE, and 2-D PAGE
> Western Blot and ELISA
> Functional Analysis
> Endotoxin and contaminating DNA measurement
> Analytical SEC and IEC
> Glycosylation Analysis
> N-terminal Sequencing and Amino Acid Analysis
> Peptide mapping by HPLC and Mass-Spec