Crystal structure of the RUN domain of the RAP2-interacting protein x.
Rap2-interacting protein x (RPIPx) is a homolog of RPIP8, a specific effector of Rap2 GTPase. The N-terminal region of RPIP8, which contains the RUN domain, interacts with Rap2. Using cell-free synthesis and NMR, we determined that the region encompassing residues 83-255 of mouse RPIPx, which is 40-residues larger than the predicted RUN domain (residues 113-245), is the minimum fragment that forms a correctly folded protein. This fragment, the RPIPx RUN domain, interacted specifically with Rap2B in vitro in a nucleotide-dependent manner. The crystal structure of the RPIPx RUN domain was determined at 2.0 A of resolution by the multiwavelength anomalous dispersion (MAD) method. The RPIPx RUN domain comprises eight anti-parallel alpha-helices, which form an extensive hydrophobic core, followed by an extended segment. The residues in the core region are highly conserved, suggesting the conservation of the RUN domain-fold among the RUN domain-containing proteins. The residues forming a positively charged surface are conserved between RPIP8 and its homologs, suggesting that this surface is important for Rap2 binding. In the crystal the putative Rap2 binding site of the RPIPx RUN domain interacts with the extended segment in a segment-swapping manner.
Kukimoto-Niino
Mutsuko
M
RIKEN Genomic Sciences Center, Yokohama 230-0045, Japan.
Takagi
Tetsuo
T
Akasaka
Ryogo
R
Murayama
Kazutaka
K
Uchikubo-Kamo
Tomomi
T
Terada
Takaho
T
Inoue
Makoto
M
Watanabe
Satoru
S
Tanaka
Akiko
A
Hayashizaki
Yoshihide
Y
Kigawa
Takanori
T
Shirouzu
Mikako
M
Yokoyama
Shigeyuki
S
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry*; D018360:Crystallography, X-Ray; D018076:DNA, Complementary / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D020668:rap GTP-Binding Proteins / Q000737:chemistry
D002352:Carrier Proteins; D018076:DNA, Complementary; D047908:Intracellular Signaling Peptides and Proteins; C486394:Rap2ip protein, mouse; C492137:Rap2a protein, mouse; D020668:rap GTP-Binding Proteins