The biochemical effect of Ser167 phosphorylation on Chlamydomonas reinhardtii centrin.
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.
Department of Biochemistry, Center for Structural Biology, 5140 BIOSCI/MRBIII, Vanderbilt University, Nashville, TN 37232-8725, USA.
Susan M
SM
Meyn
Christina
C
Seda
Muriel
M
Campbell
Kevin L
KL
Weiss
Haitao
H
Hu
Belinda
B
Pastrana-Rios
Walter J
WJ
Chazin
D000818:Animals; D002135:Calcium-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016825:Chlamydomonas reinhardtii / Q000235:genetics / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular; D010766:Phosphorylation; D010768:Phosphoserine / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D055672:Static Electricity
D002135:Calcium-Binding Proteins; D010768:Phosphoserine; D017868:Cyclic AMP-Dependent Protein Kinases