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Close proximity of phosphorylation sites to ligand in the phosphoproteome of the extreme thermophile T hermus thermophilus HB 8
Author(s) -
Takahata Yoshio,
Inoue Masao,
Kim Kwang,
Iio Yota,
Miyamoto Masaaki,
Masui Ryoji,
Ishihama Yasushi,
Kuramitsu Seiki
Publication year - 2012
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.201100573
Subject(s) - phosphorylation , protein phosphorylation , biochemistry , ligand (biochemistry) , protein subunit , biology , serine , threonine , cofactor , thermophile , eubacterium , substrate level phosphorylation , chemistry , enzyme , protein kinase a , receptor , bacteria , genetics , gene
We performed phosphoproteome analysis of proteins from the extremely thermophilic G ram‐negative eubacterium T hermus thermophilus HB 8 using gel‐free mass spectrometric method. We identified 52 phosphopeptides from 48 proteins and determined 46 phosphorylation sites: 30 on serine, 12 on threonine, and 4 on tyrosine. The identified phosphoproteins are known to be involved in a wide variety of cellular processes. To help elucidate the functional roles of these phosphorylation events, we mapped the phosphorylation sites on the known tertiary structures of the respective proteins. In all, we succeeded in mapping 46 sites (approximately 88%) on the corresponding structures. Most of the phosphorylation sites were found to be located on loops and terminal regions of the secondary structures. Surprisingly, 28 of these sites were situated at or near the active site of the enzyme. In particular, 18 sites were within 4 Å of the ligand, including substrate or cofactor. Such structural locations suggest direct effects of the phosphorylation on the binding of ligand in addition to inducing a conformational change. Interestingly, 19 of these 28 phosphorylation sites were situated near the phosphate moiety of a substrate or cofactor. In oligomeric proteins, 5 phosphorylation sites were found at the subunit interface. Based on these results, we propose a regulatory mechanism that involves S er/ T hr/ T yr phosphorylation in T . thermophilus HB 8.