| 1. |
- Nore, Beston F, et al.
(författare)
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Identification of phosphorylation sites within the SH3 domains of Tec family tyrosine kinases
- 2003
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Ingår i: Biochimica et Biophysica Acta. - 0006-3002 .- 1878-2434. ; 1645:2, s. 123-132
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Tidskriftsartikel (refereegranskat)abstract
- Tec family protein tyrosine kinases (TFKs) play a central role in hematopoietic cellular signaling. Initial activation takes place through specific tyrosine phosphorylation situated in the activation loop. Further activation occurs within the SH3 domain via a transphosphorylation mechanism, which for Bruton's tyrosine kinase (Btk) affects tyrosine 223. We found that TFKs phosphorylate preferentially their own SH3 domains, but differentially phosphorylate other member family SH3 domains, whereas non-related SH3 domains are not phosphorylated. We demonstrate that SH3 domains are good and reliable substrates. We observe that transphosphorylation is selective not only for SH3 domains, but also for dual SH3SH2 domains. However, the dual domain is phosphorylated more effectively. The major phosphorylation sites were identified as conserved tyrosines, for Itk Y180 and for Bmx Y215, both sites being homologous to the Y223 site in Btk. There is, however, one exception because the Tec-SH3 domain is phosphorylated at a non-homologous site, nevertheless a conserved tyrosine, Y206. Consistent with these findings, the 3D structures for SH3 domains point out that these phosphorylated tyrosines are located on the ligand-binding surface. Because a number of Tec family kinases are coexpressed in cells, it is possible that they could regulate the activity of each other through transphosphorylation.
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| 2. |
- Westerlind, Ulrika, et al.
(författare)
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Ligands of the asialoglycoprotein receptor for targeted gene delivery, part 1: Synthesis of and binding studies with biotinylated cluster glycosides containing N-acetylgalactosamine.
- 2004
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Ingår i: Glycoconjugate journal. - : Springer. - 0282-0080 .- 1573-4986. ; 21:5, s. 227-41
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Tidskriftsartikel (refereegranskat)abstract
- In order to develop the non-viral Bioplex vector system for targeted delivery of genes to hepatocytes, we have evaluated the structure-function relationship for a number of synthetic ligands designed for specific interaction with the hepatic lectin ASGPr. Biotinylated ligand derivatives containing two, three or six beta-linked N-acetylgalactosamine (GalNAc) residues were synthesized, bound to fluorescent-labeled streptavidin and tested for binding and uptake to HepG2 cells using flow cytometry analysis (FACS). Uptake efficiency increased with number of displayed GalNAc units per ligand, in a receptor dependent manner. Thus, a derivative displaying six GalNAc units showed the highest uptake efficacy both in terms of number of internalizing cells and increased amount of material taken up per each cell. However, this higher efficiency was shown to be due not so much to higher number of sugar units, but to higher accessibility of the sugar units for interaction with the receptor (longer spacer). Improving the flexibility and accessibility of a trimeric GalNAc ligand through use of a longer spacer markedly influenced the uptake efficiency, while increasing the number of GalNAc units per ligand above three only provided a minor contribution to the overall affinity. We hereby report the details of the chemical synthesis of the ligands and the structure-function studies in vitro.
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