ProteoMod: A new tool to quantitate protein post-translational modifications.
Post-translational modifications (PTMs) are known to regulate biological processes by controlling protein function. The effect of a PTM on protein function depends critically on the position and the number of modifications. While there are convenient methods available to qualitatively examine modifications like phosphorylation, glycosylation, acetylation and methylation, methods available for their quantitative assessment are cumbersome. We have developed a new tool that allows quantitation of the number of phosphorylation events in proteins with ease. The "ProteoMod" tool depends on shifts in the isoelectric points of proteins upon post-translational change. The extent of shift exhibited upon phosphorylation is algorithmically converted into the number of phosphorylations conferred. The validity of ProteoMod was confirmed by examining proteins with previously known number of phosphorylations. The list of proteins examined included HSP27, HSP70 and tumor suppressor p53. The approach can also be applied to estimate modifications like acetylation, methylation and sialylation in proteins. We analyzed shifts in isoelectric points due to sialylation events in N-glycoproteins. Using influenza hemagglutinin we show that shifts in isoelectric points correlate with intracellular distribution of this model membrane protein. In addition to extending the application of two dimensional gel electrophoresis to quantitate modifications, our study also highlights its potential use in cell biology.
Kumar
Yadunanda
Y
Department of Biochemistry, Indian Institute of Science, Bangalore, India.
Khachane
Amit
A
Belwal
Mukta
M
Das
Sanjeev
S
Somsundaram
Kumar
K
Tatu
Utpal
U
D000465:Algorithms; D018840:HSP70 Heat-Shock Proteins / Q000737:chemistry / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000737:chemistry / Q000378:metabolism; D007526:Isoelectric Point; D008970:Molecular Weight; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D015203:Reproducibility of Results
D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D011506:Proteins