Dissertation
STRUCTURAL ANALYSIS AND REGULATORY MECHANISMS OF CALCIUM/CALMODULIN-DEPENDENT PROTEIN KINASE (CCAMK)
Doctor of Philosophy (PhD), Washington State University
01/2012
Handle:
https://hdl.handle.net/2376/111594
Abstract
Chimeric calcium/calmodulin-dependent protein kinase (CCaMK) is a plant-specific serine/threonine protein kinase carrying unique structural features consisting of an N-terminal catalytic domain, a calmodulin-binding domain and a C-terminal visinin-like Ca+2-binding domain, with a dual regulatory mechanism involving calcium and calcium-bound calmodulin. The binding of calcium to the visinin domain of CCaMK results in the autophosphorylation at T271. This increases its affinity to calmodulin and a subsequent increase in its substrate phosphorylation capacity. CCaMK is found to be the "master switch" in decoding the calcium signal during plant-microbe interactions, and the autophosphorylation of DMI3/CCaMK plays a central role in the establishment of both root nodule and mycorrhizal symbioses.
Using LC/MS-MS analysis we identified two additional autophosphorylation sites in DMI3/CCaMK, which are phosphorylated in calcium-dependent manner. Using biochemical approaches, we found that the autophosphorylation of CCaMK at these residues can change both its interaction with calmodulin and control the kinase activity. The autophosphorylation of one of the newly-identified residues in the calmodulin binding domain is proven to be critical for both root nodule and mycorrhizal symbioses.
The autoinhibitory/calmodulin-binding domain is believed to be responsible for the inactivation of CCaMK in the absence of calcium and calmodulin. Further studies on intra-molecular interaction between the autoinhibitory domain and catalytic domain of CCaMK identified several residues responsible for this inhibitory mechanism.
These results document the existence of a calcium/calmodulin-dependent self-regulatory mechanism during plant: microbe symbioses. Further research should help in understanding this phenomena and transfer this know how to non-leguminous crops to increase crop production and decrease world hunger.
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Details
- Title
- STRUCTURAL ANALYSIS AND REGULATORY MECHANISMS OF CALCIUM/CALMODULIN-DEPENDENT PROTEIN KINASE (CCAMK)
- Creators
- Pratyush Routray
- Contributors
- B.W. Poovaiah (Advisor)Raymond Reeves (Committee Member)Patricia Okubara (Committee Member)Amit Dhingra (Committee Member)
- Awarding Institution
- Washington State University
- Academic Unit
- Program in Molecular Plant Sciences
- Theses and Dissertations
- Doctor of Philosophy (PhD), Washington State University
- Number of pages
- 124
- Identifiers
- 99900581857701842
- Language
- English
- Resource Type
- Dissertation