Differential steady-state tyrosine phosphorylation of two oligomeric forms of mitochondrial F0F1ATPsynthase:: A structural proteomic analysis

被引：29

作者：

Di Pancrazio, F ^{[1
]}

Bisetto, E ^{[1
]}

Alverdi, V ^{[1
]}

Mavelli, I ^{[1
]}

Esposito, G ^{[1
]}

Lippe, G ^{[1
]}

机构：

[1] Univ Udine, Dept Biomed Sci & Technol, MATI Ctr Excellence, I-33100 Udine, Italy

来源：

PROTEOMICS | 2006年 / 6卷 / 03期

关键词：

blue native (BN)-PAGE; dimerization; mitochondrial F(0)F(1)ATPsynthase; LC-ESI/MS; phosphotyrosines;

D O I：

10.1002/pmic.200500077

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

We investigated tyrosine phosphorylation of F(0)F(1)ATPsynthase using 3-D blue native (BN)-SDS-PAGE, a refinement of the electrophoretic analysis of mitochondrial complexes. Bovine heart mitochondria were detergent-solubilized and subjected to BN-PAGE. Bands of ATPsynthase monomer (Vmon) and dimer (Vdim) were excised and submitted to SDS-PAGE and immunoblotting. One protein corresponding to F-1 gamma subunit was detected by anti-phosphotyrosine antibody in monomer but not in dimer. This was confirmed by MS peptide mapping. LC-ESI/MS analysis after 3-D SDS-PAGE demonstrated phosphotyrosine in fragment 43-54. NetPhos scores predicted the phosphorylated residue to be Tyr52, in a solvent-accessible loop at the foot of the F, central stalk.

引用

页码：921 / 926

页数：6

共 44 条

[1] Determination of protein phosphorylation sites by mass spectrometry: a novel electrospray-based method [J].

Alverdi, V ;

Di Pancrazio, F ;

Lippe, G ;

Pucillo, C ;

Casetta, B ;

Esposito, G .

RAPID COMMUNICATIONS IN MASS SPECTROMETRY, 2005, 19 (22) :3343-3348

[2] Yeast mitochondrial F1F0-ATP synthase exists as a dimer:: identification of three dimer-specific subunits [J].

Arnold, I ;

Pfeiffer, K ;

Neupert, W ;

Stuart, RA ;

Schägger, H .

EMBO JOURNAL, 1998, 17 (24) :7170-7178

[3] The modulation in subunits e and g amounts of yeast ATP synthase modifies mitochondrial cristae morphology [J].

Arselin, G ;

Vaillier, J ;

Salin, B ;

Schaeffer, J ;

Giraud, MF ;

Dautant, A ;

Brèthes, D ;

Velours, J .

JOURNAL OF BIOLOGICAL CHEMISTRY, 2004, 279 (39) :40392-40399

[4] Ca2+-modulated phosphorylation of a low-molecular-mass polypeptide in rat liver mitochondria:: Evidence that it is identical with subunit c of F0F1-ATPase [J].

Azarashvily, TS ;

Tyynelä, J ;

Baumann, M ;

Evtodienko, YV ;

Saris, NEL .

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 2000, 270 (03) :741-744

[5]

BLAIR PV, 1967, METHOD ENZYMOL, V10, P75

[6] Sequence and structure-based prediction of eukaryotic protein phosphorylation sites [J].

Blom, N ;

Gammeltoft, S ;

Brunak, S .

JOURNAL OF MOLECULAR BIOLOGY, 1999, 294 (05) :1351-1362

[7] Phosphorylation of Y845 on the epidermal growth factor receptor mediates binding to the mitochondrial protein cytochrome c oxidase subunit II [J].

Boerner, JL ;

Demory, ML ;

Silva, C ;

Parsons, SJ .

MOLECULAR AND CELLULAR BIOLOGY, 2004, 24 (16) :7059-7071

[8]

BRADFORD MM, 1976, ANAL BIOCHEM, V72, P248, DOI 10.1016/0003-2697(76)90527-3

[9]

Brookes PS, 2002, PROTEOMICS, V2, P969, DOI 10.1002/1615-9861(200208)2:8<969::AID-PROT969>3.0.CO

[10]

2-3

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