Summary of Research Project Results Under the JSPS FY2000
"Research for the future Program"



1.Research Institution University of Tsukuba
 
2.Research Area Life Sciences
 
3.Research Field Structure and Functional Control Mechanism of Biomolecules
 
4.Term of Project FY1996〜FY2000
 
5.Project Number 96L00501
 
6.Title of Project Structural Biology of Proteins Involved in Cellular Signal Transduction

7.Projetct Leader
Name Institution,Department Title of Position
Toshiyuki, Tanaka Institute of Applied Biochemistry Associate Professor

8.Core Members

Names Institution,Department Title of Position
Mitsuhiko, Ikura Ontario Cancer Institute, University of Toronto Professor

9.Cooperating Researchers

10.Summary of Research Results

1. Structure Determination of Proteins Involved in Signal Transduction
(1) Calcium Signal Transduction
We have determined the solution structure of calcium-bound calmodulin (CaM) complexed with is potent antagonist W-7 and revealed the mechanism by which W-7 inhibits CaM to activate the CaM-dependent enzymes.
Retinal recoverin contains a covalently attached myristoyl or related acyl group at its N-terminus. Ca2+ binding to myristoylated, not unmyristoylated, recombinant recoverin induces the translocation of the protein from cytosol to membranes, suggesting that the myristoyl group is crucial for membrane targeting. We have determined the solution structures of recoverin in the Ca2+-free and -bound states and revealed that Ca2+ induces the unclamping and extrusion of the myristoyl group, which is sequestered in a deep hydrophobic pocket of the protein in the Ca2+-free state, enabling it to interact with a lipid bilayer membrane.
(2) His-Asp Phosphorelay Signal Transduction
We have determined the solution structures of the homodimeric core domain and the kinase domain of EnvZ and revealed its signal transduction mechanism through dimerization.
(3) Transcription Regulation
We have determined the solution structure of the DNA-binding domain of MafG and proposed a possible mechanism by which Maf family proteins recognize their consensus DNA sequences.
2. Development of "Molecular Replacement" Program
The prototype of the "Molecular Replacement" program, which was devised to do sequential backbone and side-chain assignments of mutant proteins based on the NMR data of the wild-type protein, has succeeded in complete assignments of several mutant proteins with a single amino acid substitution. Development of a program to achieve NOE assignments is in progress.

11.Key Words

(1)Signal Transduction、(2)Protein、(3)Solution Structure
(4)NMR、(5)Calmodulin、(6)Recoverin
(7)EnvZ、(8)MafG、(9)Molecular Replacement

12.References

[Reference Articles]
Author Title of Article
J. B. Ames Molecular Mechanics of Calcium-Myristoyl Switches
Journal Volume Year Pages Concerned
Nature 389 1997 198-202

Author Title of Article
M. Osawa Solution Structure of Calmodulin-W-7 Complex: The Basis of Diversity in Molecular Recognition
Journal Volume Year Pages Concerned
J. Mol. Biol. 276 1998 165-176

Author Title of Article
T. Tanaka NMR Structure of the Histidine Kinase Domain of the E. coli Osmosensor EnvZ
Journal Volume Year Pages Concerned
Nature 396 1998 88-92

Author Title of Article
C. Tomomori Solution Structure of the Homodimeric Core Domain of Escherichia coli Histidine Kinase EnvZ
Journal Volume Year Pages Concerned
Nature Struct. Biol. 6 1999 729-734

Author Title of Article
T. Tanaka Solution Structures of C-1027 Apoprotein and Its Complex with the Aromatized Chromophore
Journal Volume Year Pages Concerned
J. Mol. Biol. 309 2001 267-283


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