Journal article

Phosphopeptide Enrichment with TiO2-Modified Membranes and Investigation of Tau Protein Phosphorylation

Yu-Jing Tan, Dexin Sui, Wei-Han Wang, Min-Hao Kuo, Gavin E Reid, Merlin L Bruening

ANALYTICAL CHEMISTRY | AMER CHEMICAL SOC | Published : 2013

Abstract

Selective enrichment of phosphopeptides prior to their analysis by mass spectrometry (MS) is vital for identifying protein phosphorylation sites involved in cellular regulation. This study describes modification of porous nylon substrates with TiO2 nanoparticles to create membranes that rapidly enrich phosphopeptides. Membranes with a 22-mm diameter bind 540 nmol of phosphoangiotensin and recover 70% of the phosphopeptides in mixtures with a 15-fold excess of nonphosphorylated proteins. Recovery is 90% for a pure phosphopeptide. Insertion of small membrane disks into HPLC fittings allows rapid enrichment from 5 mL of 1 fmol/μL phosphoprotein digests and concentration into small-volume (tens ..

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University of Melbourne Researchers

Grants

Awarded by U.S. National Institutes of Health


Awarded by National Science Foundation


Awarded by NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES


Awarded by NATIONAL INSTITUTE ON AGING


Funding Acknowledgements

We are grateful to the U.S. National Institutes of Health (GM080511 to M.B. and AG039768 to M.H.K.) and the National Science Foundation (CHE-1152762) for funding this work. We thank Dr. Amanda Palumbo for synthesizing the H<INF>5</INF> and D<INF>5</INF> peptides, and Li Cui from the Reid Group for her assistance with phosphorylation assignment.