Forskning ved Københavns Universitet - Københavns Universitet

Forside

Lactococcus lactis provides an efficient platform for production of disulfide-rich recombinant proteins from Plasmodium falciparum

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningfagfællebedømt

BACKGROUND: The production of recombinant proteins with proper conformation, appropriate post-translational modifications in an easily scalable and cost-effective system is challenging. Lactococcus lactis has recently been identified as an efficient Gram positive cell factory for the production of recombinant protein. We and others have used this expression host for the production of selected malaria vaccine candidates. The safety of this production system has been confirmed in multiple clinical trials. Here we have explored L. lactis cell factories for the production of 31 representative Plasmodium falciparum antigens with varying sizes (ranging from 9 to 90 kDa) and varying degree of predicted structural complexities including eleven antigens with multiple predicted structural disulfide bonds, those which are considered difficult-to-produce proteins.

RESULTS: Of the 31 recombinant constructs attempted in the L. lactis expression system, the initial expression efficiency was 55% with 17 out of 31 recombinant gene constructs producing high levels of secreted recombinant protein. The majority of the constructs which failed to produce a recombinant protein were found to consist of multiple intra-molecular disulfide-bonds. We found that these disulfide-rich constructs could be produced in high yields when genetically fused to an intrinsically disorder protein domain (GLURP-R0). By exploiting the distinct biophysical and structural properties of the intrinsically disordered protein region we developed a simple heat-based strategy for fast purification of the disulfide-rich protein domains in yields ranging from 1 to 40 mg/l.

CONCLUSIONS: A novel procedure for the production and purification of disulfide-rich recombinant proteins in L. lactis is described.

OriginalsprogEngelsk
Artikelnummer55
TidsskriftMicrobial Cell Factories
Vol/bind17
Antal sider13
ISSN1475-2859
DOI
StatusUdgivet - 2018

Antal downloads er baseret på statistik fra Google Scholar og www.ku.dk


Ingen data tilgængelig

ID: 199300265