2012
DOI: 10.1016/j.pep.2011.11.016
|View full text |Cite
|
Sign up to set email alerts
|

Abstract: Affinity purification is a useful approach for purification of recombinant proteins. Eukaryotic expression systems have become more frequently used at the expense of prokaryotic systems since they afford recombinant eukaryotic proteins with post-translational modifications similar or identical to the native ones.Here, we present a one-step affinity purification set-up suitable for the purification of secreted proteins. The set-up is based on the interaction between biotin and mutated streptavidin. Drosophila S… Show more

Help me understand this report

Search citation statements

Order By: Relevance

Paper Sections

Select...
2
1
1
1

Citation Types

2
66
0

Year Published

2013
2013
2017
2017

Publication Types

Select...
8

Relationship

1
7

Authors

Journals

citations
Cited by 37 publications
(68 citation statements)
references
References 39 publications
2
66
0
Order By: Relevance
“…Then, the sequence corresponding to the extracellular part of mouse GCPII (amino acids 45‐752) was amplified by PCR using primers 5′‐aaaagatctaaaccttccaatgaagctactgg‐3′ and 5′‐aaactcgagttaagctacttccctcagagtc‐3′ (restriction sites introduced into the sequence are underlined; the primers introduced a Bgl II site at the 5′ end and an Xho I site at the 3′ end). The resulting DNA fragment was cleaved with Bgl II and Xho I and ligated into pMT/BiP/AviTEV/rhGCPII plasmid cleaved with the same endonucleases. The correct sequence of the resulting plasmid pMT/BiP/Avi‐mGCPII was verified by DNA sequencing.…”
Section: Methodsmentioning
confidence: 99%
“…Then, the sequence corresponding to the extracellular part of mouse GCPII (amino acids 45‐752) was amplified by PCR using primers 5′‐aaaagatctaaaccttccaatgaagctactgg‐3′ and 5′‐aaactcgagttaagctacttccctcagagtc‐3′ (restriction sites introduced into the sequence are underlined; the primers introduced a Bgl II site at the 5′ end and an Xho I site at the 3′ end). The resulting DNA fragment was cleaved with Bgl II and Xho I and ligated into pMT/BiP/AviTEV/rhGCPII plasmid cleaved with the same endonucleases. The correct sequence of the resulting plasmid pMT/BiP/Avi‐mGCPII was verified by DNA sequencing.…”
Section: Methodsmentioning
confidence: 99%
“…To further investigate these differences, we expressed different versions of recombinant AMP1 protein to test its catalytic activity against the HsGCPII substrates NAAG and folyl-poly-g-L-glutamic acids. However, we were not able to produce sufficient amounts of AMP1 protein by using either Escherichia coli or the insect cell expression system, which are routinely used for recombinant GCPII production (Tykvart et al, 2012). As an alternative approach to test for functional conservation between the two proteins, we expressed a MYC-tagged version of HsGCPII under control of the CaMV 35S promoter in amp1-1.…”
Section: No Evidence For Functional Conservation Between Amp1 and Hsgmentioning
confidence: 99%
“…Expression and Purification of Avi-NaalL and AviNaalL(E416A)-Preparation and purification of Avi-NaalL was performed as described previously (5). Briefly, Drosophila S2 cells expressing E. coli biotin-ligase (BirA) in the endoplasmic reticulum were transfected with pMT/BiP/AviTEV/rhNaalL (encoding the extracellular portion (aa 28 -740) of NAALADase L with an N-terminal Avi tag and TEV protease cleavage site.…”
Section: Methodsmentioning
confidence: 99%
“…recombinant GCPII preparation (Avi-GCPII) and its catalytically ineffective E424A mutant as a corresponding control pair (5,25).…”
Section: Enzymatic Activities Of Naaladase Lmentioning
confidence: 99%