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Immunoaffinity isolation of Btk´s signalosome, a proteomic approach to identifying interacting proteins
Södertörn University College, School of Life Sciences.
2006 (English)Independent thesis Basic level (professional degree), 10 points / 15 hpStudent thesis
Abstract [en]

The Signalosome is a term used to define a putative signalling complex, which assembles near the plasma membrane in response to external signals received at cell surface receptors and then migrates towards downstream effectors. It is proposed to regulate the level of intracellular Ca2+ and subsequent downstream signalling events. To date it has been defined to consist of BTK, BLNK, BCAP, VAV, PLCγ2 and PI3K1-4 in B-Cells.

This work entailed initiating a new proteomic approach to investigate the nature and extent of Bruton’s tyrosine kinase, Btk, involvement in the signalosome – inherently, the aim was to study multiple interactions of Btk with other molecules. By transfecting host cells with a Btk gene-transfer plasmid, virus particles were produced that were used to up-regulate and analyse the expression of Btk in three haematopoietic cell lines: B-cells, Pre-B-cells and a myeloid cancer cell. The construction of a new gene-transfer vector was successfully carried out by plasmid sub-cloning and it was subsequently found to effectively transfect the host cells and produce virus particles. The recombinant virus particles were employed with success in transducing three haematopoietic cell lines and with immunopurification and subsequent gel separation protein signalosome complexes were obtained ready for analysis by mass spectrometrical fingerprinting (to be carried out as a joint effort in Mount Sinai Hospital in Toronto, Canada).

Place, publisher, year, edition, pages
Huddinge: Institutionen för livsvetenskaper , 2006. , 19 p.
Keyword [en]
Btk, signalosome, interacting proteins, gene therapy
National Category
Biochemistry and Molecular Biology
Identifiers
URN: urn:nbn:se:sh:diva-1114OAI: oai:DiVA.org:sh-1114DiVA: diva2:15167
Uppsok
bio-/geovetenskap
Supervisors
Examiners
Available from: 2007-03-27 Created: 2007-03-27

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CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf