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Defining the physiological role of phosphatidylinositol 3-kinase protein kinase activity

Foukas, Lazaros; (2003) Defining the physiological role of phosphatidylinositol 3-kinase protein kinase activity. Doctoral thesis (Ph.D), UCL (University College London). Green open access

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Abstract

Class I PI3-kinases possess an intrinsic protein kinase activity in addition to their lipid kinase activity. The best characterised target of this activity is the Ser608 of the p85 regulatory subunit in the case of the α isoform of the p110 catalytic subunit or the catalytic subunit itself, in the case of the β, γ and δ isoforms, respectively. Furthermore, it has been shown that Ser608 phosphorylation results in downregulation of the lipid kinase activity of the enzyme in vitro. The implications of this finding are very important, as it could provide a mechanism for the regulation of the lipid kinase activity of the enzyme in vivo. By generating phosphospecific antibodies against Ser608, we have been able to demonstrate that Ser608 can be phosphorylated in vivo. We have also showed that mutation of Ser608 to either alanine or glutamic acid interferes with the integrity of the p85α/p110α heterodimer and this likely explains the reduced lipid kinase activity associated with S608 phosphorylation. Also, we have demonstrated that certain class I PI3-kinases phosphorylate novel exogenous substrates, namely the translational regulator 4EBP1 and the small GTPase H-Ras in vitro. In an effort to discriminate between the two activities we have employed two different approaches. First, we have found that certain methylxanthines inhibit the lipid and the protein kinase activities but with very dinstict potencies. Also, a panel of synthetic peptides derived from class II MHC sequences activate the lipid but not the protein kinase activity of heterodimeric PI3-kinases. These findings demonstrate that each one of the two activities can be modulated independently from the other. Second, by mutating certain aminoacids in the activation loop of p110α and p110β, we have created enzyme versions that lack the lipid kinase but retain protein kinase activity. Both of these approaches could be exploited to assign specific functions to each of the lipid and protein kinase activities in cells.

Type: Thesis (Doctoral)
Qualification: Ph.D
Title: Defining the physiological role of phosphatidylinositol 3-kinase protein kinase activity
Open access status: An open access version is available from UCL Discovery
Language: English
Additional information: Thesis digitised by ProQuest.
Keywords: Biological sciences; PI3-kinase
URI: https://discovery.ucl.ac.uk/id/eprint/10107314
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