The isolation and characterisation of MHC-presented peptides from CML-derived cell-lines, with a focus on post-translational modification

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Kapoor, K.N., 2011. The isolation and characterisation of MHC-presented peptides from CML-derived cell-lines, with a focus on post-translational modification. PhD, Nottingham Trent University.

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Abstract

Phosphorylation is a key regulator of protein function and activity, and aberrant kinase activity is implicated in a wide range of malignancies, of which the bcr:abl fusion kinase found in chronic myeloid leukaemia is a classic example. As phosphopeptides are known to be presented by both the MHC class-I and class- II pathways, against which specific CD4+ and CD8+ T cell responses may be generated, study of MHC-presented phosphopeptides may reveal unique cancer antigens with direct links to the neoplastic state. Mild acid cell-surface elution is a rapid and effective method for MHC class-I peptide capture, though complicated by contamination with non-MHC peptides and poor downstream compatibility, especially with IMAC, a popular method for phosphopeptide enrichment. As an alternative to the citrate-phosphate elution buffer, a TMA-formate elution buffer is proposed. This was developed for IMAC compatibility, and osmotically balanced and supplemented to minimise cell lysis, (assessed by several assays) and used with a pH 5.5 prewash to reduce non- MHC peptide contamination. MALDI-MS/MS of MHC class-I peptides from K562- A3 cells found a sequence with high homology to a known cancer antigen as the common peak for both citrate-phosphate and TMA-formate eluted cells.

Item Type:	Thesis
Creators:	Kapoor, K.N.
Publisher:	Nottingham Trent University
Place of Publication:	Nottingham
Date:	2011
Rights:	This work is the intellectual property of the author. You may copy up to 5% of this work for private study, or personal, non-commercial research. Any reuse of the information contained within this document should be fully referenced, quoting the author, title, university, degree level and pagination. Queries or requests for any other use, or if a more substantial copy is required, should be directed in the first instance to the owner(s) of the Intellectual Property Rights.
Divisions:	Schools > School of Science and Technology
Record created by:	EPrints Services
Date Added:	09 Oct 2015 09:34
Last Modified:	09 Oct 2015 09:34
URI:	https://irep.ntu.ac.uk/id/eprint/162