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SUMMARY:Contr. talk - Small-scale expression and purification of the cytok
 ine receptor\, MPL\, using the ALiCE® cell-free system
DTSTART;VALUE=DATE-TIME:20210526T093000Z
DTEND;VALUE=DATE-TIME:20210526T095000Z
DTSTAMP;VALUE=DATE-TIME:20260722T000810Z
UID:indico-contribution-1141@lindico453.srv.lu.se
DESCRIPTION:Speakers: Julie Tucker (University of York)\nMPL (also called 
 TpoR) is the receptor for the haematopoietic cytokine\, thrombopoietin (TP
 O). Together\, MPL and TPO control the production of platelets and the mai
 ntenance of haematopoietic stem cells. Gain of function mutations in MPL c
 onstitute ˜5-10 % of driver mutations in essential\nthrombocythemia and p
 rimary myelofibrosis\, whilst loss of function mutations give rise to thro
 mbocytopenias. MPL comprises an ˜500 aa extracellular domain\, a single t
 ransmembrane helix\, and an ˜120 aa intracellular domain. The extracellul
 ar domain contains the thrombopoietin binding site\, whilst the intracellu
 lar domain provides the binding site for Janus kinase 2\, as well as other
  downstream signalling molecules\, including the Signal Transducers and Ac
 tivators of Transcription\n(STAT) proteins. Structural information on MPL 
 is lacking\, in part due to the challenges of producing recombinant protei
 n. Endogenous expression levels of MPL are very low (< 5000\nreceptors/cel
 l) and heterologous expression of full-length MPL has not been reported. H
 ere we report small-scale expression and purification of both full-length 
 and ectodomain of MPL using the ALiCE® cell-free expression system (Lenio
 Bio GmbH). This plant cell-based system allows for the expression of membr
 ane and secreted proteins within the microsomal compartment\, ensuring the
 ir correct folding\, membrane insertion and glycosylation. Optimisation of
  expression time\, plasmid\nconcentration and downstream processing allowe
 d for production and purification of glycosylated MPL\, the functional cha
 racterisation of which is ongoing.\n\nhttps://lindico453.srv.lu.se/event/2
 19/contributions/1141/
LOCATION:
URL:https://lindico453.srv.lu.se/event/219/contributions/1141/
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