In April 2016 Manchester eScholar was replaced by the University of Manchester’s new Research Information Management System, Pure. In the autumn the University’s research outputs will be available to search and browse via a new Research Portal. Until then the University’s full publication record can be accessed via a temporary portal and the old eScholar content is available to search and browse via this archive.

Multiple reaction monitoring to identify sites of protein phosphorylation with high sensitivity.

Unwin RD, Griffiths J, Leverentz M, Grallert A, Hagan IM, Whetton A

Mol Cell Proteomics. 2005;4( 8):1134-44.

Access to files

Full-text and supplementary files are not available from Manchester eScholar. Use our list of Related resources to find this item elsewhere. Alternatively, request a copy from the Library's Document supply service.

Abstract

Phosphorylation governs the activity of many proteins. Insight into molecular mechanisms in biology would be immensely improved by robust, sensitive methods for identifying precisely sites of phosphate addition. An approach to selective mapping of protein phosphorylation sites on a specific target protein of interest using LC-MS is described here. In this approach multiple reaction monitoring is used as an extremely sensitive MS survey scan for potential phosphopeptides from a known protein. This is automatically followed by peptide sequencing and subsequent location of the phosphorylation site; both of these steps occur in a single LC-MS run, providing greater efficiency of sample use. The method is capable of detecting and sequencing phosphopeptides at low femtomole levels with high selectivity. As proof of the value of this approach in an experimental setting, a key Schizosaccharomyces pombe cell cycle regulatory protein, Cyclin B, was purified, and associated proteins were identified. Phosphorylation sites on these proteins were located. The technique, which we have called multiple reaction monitoring-initiated detection and sequencing (MIDAS), is shown to be a highly sensitive approach to the determination of protein phosphorylation.

Bibliographic metadata

Type of resource:
Content type:
Publication type:
Publication form:
Published date:
Journal title:
ISSN:
Place of publication:
United States
Volume:
4( 8)
Start page:
1134
End page:
44
Pagination:
1134-44
Access state:
Active

Institutional metadata

University researcher(s):

Record metadata

Manchester eScholar ID:
uk-ac-man-scw:1d11460
Created:
29th August, 2009, 16:04:25
Last modified:
1st February, 2015, 19:05:59

Can we help?

The library chat service will be available from 11am-3pm Monday to Friday (excluding Bank Holidays). You can also email your enquiry to us.