The biochemical effect of Ser167 phosphorylation on Chlamydomonas reinhardtii centrin

Susan M. Meyn; Christina Seda; Muriel Campbell; Kevin L. Weiss; Haitao Hu; Belinda Pastrana-Rios; Walter J. Chazin

doi:10.1016/j.bbrc.2006.01.155

The biochemical effect of Ser167 phosphorylation on Chlamydomonas reinhardtii centrin

Susan M. Meyn, Christina Seda, Muriel Campbell, Kevin L. Weiss, Haitao Hu, Belinda Pastrana-Rios, Walter J. Chazin

Biological Labeling and Scattering

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13 Scopus citations

Abstract

Centrin is an EF-hand calcium-binding protein found in microtubule organizing centers of organisms ranging from algae and yeast to man. Phosphorylation in the centrin C-terminal domain occurs in mitosis and is associated with alterations in contractile fibers. To obtain insight into the structural basis for the functional effect of phosphorylation, Chlamydomonas reinhardtii centrin C-terminal domain phosphorylated at Ser167 (pCRC-C) has been produced and characterized. The structure of pCRC-C was compared to the unmodified protein by NMR spectroscopy. The effect of phosphorylation on target binding was examined for the complex of pCRC-C and a 19 residue centrin-binding fragment of Kar1. Remarkably, the efficient and selective phosphorylation by PKA was suppressed in the complex. Moreover, comparisons of NMR chemical shift differences induced by phosphorylation reveal a greater effect from phosphorylation in the context of the Kar1 complex than for the free protein. These results directly demonstrate that phosphorylation modulates the structure and biochemical activities of centrin.

Original language	English
Pages (from-to)	342-348
Number of pages	7
Journal	Biochemical and Biophysical Research Communications
Volume	342
Issue number	1
DOIs	https://doi.org/10.1016/j.bbrc.2006.01.155
State	Published - Mar 31 2006

Funding

This work was supported by the National Institutes of Health Grants RO1 GM-40120 and RO1 GM 65484 (to W.J.C.), NIH-SCORE Grant 5-S06GM08103 (to B.P.-R.), and R25 GM67072 (to Vanderbilt Summer Research Experiences for Undergraduates), with facilities supported by the Vanderbilt Center in Molecular Toxicology Grant (P50 ES00267) and Vanderbilt Ingram Cancer Center (P30 CA68485).

Funders	Funder number
NIH-SCORE	R25 GM67072, 5-S06GM08103
Vanderbilt Center in Molecular Toxicology	P50 ES00267
National Institutes of Health	RO1 GM 65484
National Institute of General Medical Sciences	R01GM040120
Vanderbilt-Ingram Cancer Center	P30 CA68485

Keywords

Calcium sensor
Calmodulin
Caltractin
Centrin
Conformational exchange
EF-hand calcium-binding protein
Energetic coupling
Kar1

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title = "The biochemical effect of Ser167 phosphorylation on Chlamydomonas reinhardtii centrin",

abstract = "Centrin is an EF-hand calcium-binding protein found in microtubule organizing centers of organisms ranging from algae and yeast to man. Phosphorylation in the centrin C-terminal domain occurs in mitosis and is associated with alterations in contractile fibers. To obtain insight into the structural basis for the functional effect of phosphorylation, Chlamydomonas reinhardtii centrin C-terminal domain phosphorylated at Ser167 (pCRC-C) has been produced and characterized. The structure of pCRC-C was compared to the unmodified protein by NMR spectroscopy. The effect of phosphorylation on target binding was examined for the complex of pCRC-C and a 19 residue centrin-binding fragment of Kar1. Remarkably, the efficient and selective phosphorylation by PKA was suppressed in the complex. Moreover, comparisons of NMR chemical shift differences induced by phosphorylation reveal a greater effect from phosphorylation in the context of the Kar1 complex than for the free protein. These results directly demonstrate that phosphorylation modulates the structure and biochemical activities of centrin.",

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AU - Meyn, Susan M.

AU - Seda, Christina

AU - Campbell, Muriel

AU - Weiss, Kevin L.

AU - Hu, Haitao

AU - Pastrana-Rios, Belinda

AU - Chazin, Walter J.

PY - 2006/3/31

Y1 - 2006/3/31

N2 - Centrin is an EF-hand calcium-binding protein found in microtubule organizing centers of organisms ranging from algae and yeast to man. Phosphorylation in the centrin C-terminal domain occurs in mitosis and is associated with alterations in contractile fibers. To obtain insight into the structural basis for the functional effect of phosphorylation, Chlamydomonas reinhardtii centrin C-terminal domain phosphorylated at Ser167 (pCRC-C) has been produced and characterized. The structure of pCRC-C was compared to the unmodified protein by NMR spectroscopy. The effect of phosphorylation on target binding was examined for the complex of pCRC-C and a 19 residue centrin-binding fragment of Kar1. Remarkably, the efficient and selective phosphorylation by PKA was suppressed in the complex. Moreover, comparisons of NMR chemical shift differences induced by phosphorylation reveal a greater effect from phosphorylation in the context of the Kar1 complex than for the free protein. These results directly demonstrate that phosphorylation modulates the structure and biochemical activities of centrin.

AB - Centrin is an EF-hand calcium-binding protein found in microtubule organizing centers of organisms ranging from algae and yeast to man. Phosphorylation in the centrin C-terminal domain occurs in mitosis and is associated with alterations in contractile fibers. To obtain insight into the structural basis for the functional effect of phosphorylation, Chlamydomonas reinhardtii centrin C-terminal domain phosphorylated at Ser167 (pCRC-C) has been produced and characterized. The structure of pCRC-C was compared to the unmodified protein by NMR spectroscopy. The effect of phosphorylation on target binding was examined for the complex of pCRC-C and a 19 residue centrin-binding fragment of Kar1. Remarkably, the efficient and selective phosphorylation by PKA was suppressed in the complex. Moreover, comparisons of NMR chemical shift differences induced by phosphorylation reveal a greater effect from phosphorylation in the context of the Kar1 complex than for the free protein. These results directly demonstrate that phosphorylation modulates the structure and biochemical activities of centrin.

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KW - Caltractin

KW - Centrin

KW - Conformational exchange

KW - EF-hand calcium-binding protein

KW - Energetic coupling

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The biochemical effect of Ser167 phosphorylation on Chlamydomonas reinhardtii centrin

Abstract

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Keywords

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