A surface-activated chemical ionization approach allows quantitative phosphorylation analysis of the cyclin-dependent kinase inhibitor Sic1 phosphorylated on Ser201.
Rapid Commun Mass Spectrom
; 26(13): 1527-32, 2012 Jul 15.
Article
in En
| MEDLINE
| ID: mdl-22638969
ABSTRACT
RATIONALE Quantitative phosphoproteomics represents a front line for functional proteomics and hence for systems biology. Here we present a new application of the surface-activated chemical ionization (SACI) technology for quantitative phosphoproteomics analysis. The main advantages of SACI-MS technology are high sensitivity, quantitative accuracy and matrix effect reduction, which allow quantitative estimations. METHODS:
A SACI-MS approach was used to investigate the quantitative in vivo phosphorylation of the cyclin-dependent kinase inhibitor Sic1, a low-abundance protein of Saccharomyces cerevisiae, which is phosphorylated on Ser201 by casein kinase 2 (CK2) and compared its phosphorylation status in cells growing in two different carbon sources (glucose or ethanol).RESULTS:
Our relative quantification indicated that the Sic1-Ser201 phosphorylation level is about 2-fold higher in ethanol- than in glucose-growing cells, proportional to the Sic1 protein level. This finding is coherent with results of western blot analysis using anti-phospho-Ser201-specific antibody, validating the results obtained with this new SACI approach.CONCLUSIONS:
The findings presented in this paper indicate that the innovative LC/SACI-MS method, coupled with immunoprecipitation, is a powerful device to obtain quantitative information on the phosphorylation state of low abundance proteins.
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Serine
/
Mass Spectrometry
/
Saccharomyces cerevisiae Proteins
/
Cyclin-Dependent Kinase Inhibitor Proteins
Language:
En
Journal:
Rapid Commun Mass Spectrom
Year:
2012
Document type:
Article
Affiliation country:
Italia