PDBsum entry 1ioz

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protein ligands links
Signaling protein PDB id
Jmol PyMol
Protein chain
162 a.a. *
Waters ×180
* Residue conservation analysis
PDB id:
Name: Signaling protein
Title: Crystal structure of thE C-ha-ras protein prepared by the cell-free synthesis
Structure: Transforming protein p21/h-ras-1. Chain: a. Fragment: residues 1-171. Synonym: c-h-ras. Engineered: yes
Source: Homo sapiens. Human. Organism_taxid: 9606. Other_details: cell-free protein synthesis system
Biol. unit: Dimer (from PQS)
2.00Å     R-factor:   0.238     R-free:   0.283
Authors: T.Kigawa,E.Yamaguchi-Nunokawa,K.Kodama,T.Matsuda,T.Yabuki, Riken Structural Genomics/proteomics Initiative (Rsgi)
Key ref: T.Kigawa et al. (2002). Selenomethionine incorporation into a protein by cell-free synthesis. J Struct Funct Genomics, 2, 29-35. PubMed id: 12836672
18-Apr-01     Release date:   03-Oct-01    
Go to PROCHECK summary

Protein chain
Pfam   ArchSchema ?
P01112  (RASH_HUMAN) -  GTPase HRas
189 a.a.
162 a.a.
Key:    PfamA domain  Secondary structure  CATH domain

 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     membrane   9 terms 
  Biological process     response to isolation stress   51 terms 
  Biochemical function     nucleotide binding     6 terms  


J Struct Funct Genomics 2:29-35 (2002)
PubMed id: 12836672  
Selenomethionine incorporation into a protein by cell-free synthesis.
T.Kigawa, E.Yamaguchi-Nunokawa, K.Kodama, T.Matsuda, T.Yabuki, N.Matsuda, R.Ishitani, O.Nureki, S.Yokoyama.
Multi-wavelength anomalous diffraction phasing is especially useful for high-throughput structure determinations. Selenomethionine substituted proteins are commonly used for this purpose. However, the cytotoxicity of selenomethionine drastically reduces the efficiency of its incorporation in in vivo expression systems. In the present study, an improved E. coli cell-free protein synthesis system was used to incorporate selenomethionine into a protein, so that highly efficient incorporation could be achieved. A milligram quantity of selenomethionine-containing Ras was obtained using the cell-free system with dialysis. The mass spectrometry analysis showed that more than 95% of the methionine residues were substituted with selenomethionine. The crystal of this protein grew under the same conditions and had the same unit cell constants as those of the native Ras protein. The three-dimensional structure of this protein, determined by multi-wavelength anomalous diffraction phasing, was almost the same as that of the Ras protein prepared by in vivo expression. Therefore, the cell-free synthesis system could become a powerful protein expression method for high-throughput structure determinations by X-ray crystallography.

Literature references that cite this PDB file's key reference

  PubMed id Reference
21103555 D.J.Stigers, Z.I.Watts, J.E.Hennessy, H.K.Kim, R.Martini, M.C.Taylor, K.Ozawa, J.W.Keillor, N.E.Dixon, and C.J.Easton (2011).
Incorporation of chlorinated analogues of aliphatic amino acids during cell-free protein synthesis.
  Chem Commun (Camb), 47, 1839-1841.  
20637904 F.Junge, S.Haberstock, C.Roos, S.Stefer, D.Proverbio, V.Dötsch, and F.Bernhard (2011).
Advances in cell-free protein synthesis for the functional and structural analysis of membrane proteins.
  N Biotechnol, 28, 262-271.  
20949621 L.Gremer, T.Merbitz-Zahradnik, R.Dvorsky, I.C.Cirstea, C.P.Kratz, M.Zenker, A.Wittinghofer, and M.R.Ahmadian (2011).
Germline KRAS mutations cause aberrant biochemical and physical properties leading to developmental disorders.
  Hum Mutat, 32, 33-43.  
20382987 H.Walden (2010).
Selenium incorporation using recombinant techniques.
  Acta Crystallogr D Biol Crystallogr, 66, 352-357.  
18781689 A.R.Goerke, and J.R.Swartz (2009).
High-level cell-free synthesis yields of proteins containing site-specific non-natural amino acids.
  Biotechnol Bioeng, 102, 400-416.  
19801550 K.Murayama, S.Nakayama, M.Kato-Murayama, R.Akasaka, N.Ohbayashi, Y.Kamewari-Hayami, T.Terada, M.Shirouzu, T.Tsurumi, and S.Yokoyama (2009).
Crystal structure of epstein-barr virus DNA polymerase processivity factor BMRF1.
  J Biol Chem, 284, 35896-35905.
PDB code: 2z0l
17846751 Y.Ogra, T.Kitaguchi, N.Suzuki, and K.T.Suzuki (2008).
In vitro translation with [34S]-labeled methionine, selenomethionine, and telluromethionine.
  Anal Bioanal Chem, 390, 45-51.  
17660253 C.N.Cronin, K.B.Lim, and J.Rogers (2007).
Production of selenomethionyl-derivatized proteins in baculovirus-infected insect cells.
  Protein Sci, 16, 2023-2029.  
17456747 D.F.Savage, C.L.Anderson, Y.Robles-Colmenares, Z.E.Newby, and R.M.Stroud (2007).
Cell-free complements in vivo expression of the E. coli membrane proteome.
  Protein Sci, 16, 966-976.  
17190834 K.Murayama, M.Shirouzu, Y.Kawasaki, M.Kato-Murayama, K.Hanawa-Suetsugu, A.Sakamoto, Y.Katsura, A.Suenaga, M.Toyama, T.Terada, M.Taiji, T.Akiyama, and S.Yokoyama (2007).
Crystal structure of the rac activator, Asef, reveals its autoinhibitory mechanism.
  J Biol Chem, 282, 4238-4242.
PDB code: 2dx1
17348022 M.Abe, S.Ohno, T.Yokogawa, T.Nakanishi, F.Arisaka, T.Hosoya, T.Hiramatsu, M.Suzuki, T.Ogasawara, T.Sawasaki, K.Nishikawa, M.Kitamura, H.Hori, and Y.Endo (2007).
Detection of structural changes in a cofactor binding protein by using a wheat germ cell-free protein synthesis system coupled with unnatural amino acid probing.
  Proteins, 67, 643-652.  
17288516 M.Forstner, L.Leder, and L.M.Mayr (2007).
Optimization of protein expression systems for modern drug discovery.
  Expert Rev Proteomics, 4, 67-78.  
17947230 S.K.Olsen, N.Ota, S.Kishishita, M.Kukimoto-Niino, K.Murayama, H.Uchiyama, M.Toyama, T.Terada, M.Shirouzu, O.Kanagawa, and S.Yokoyama (2007).
Crystal Structure of the Interleukin-15{middle dot}Interleukin-15 Receptor {alpha} Complex: INSIGHTS INTO TRANS AND CIS PRESENTATION.
  J Biol Chem, 282, 37191-37204.
PDB code: 2psm
17295314 S.Tomić, B.Bertosa, T.Wang, and R.C.Wade (2007).
COMBINE analysis of the specificity of binding of Ras proteins to their effectors.
  Proteins, 67, 435-447.  
18167031 T.Yabuki, Y.Motoda, K.Hanada, E.Nunokawa, M.Saito, E.Seki, M.Inoue, T.Kigawa, and S.Yokoyama (2007).
A robust two-step PCR method of template DNA production for high-throughput cell-free protein synthesis.
  J Struct Funct Genomics, 8, 173-191.  
17020876 M.Suzuki, R.Roy, H.Zheng, N.Woychik, and M.Inouye (2006).
Bacterial bioreactors for high yield production of recombinant protein.
  J Biol Chem, 281, 37559-37565.  
16703415 N.Chumpolkulwong, K.Sakamoto, A.Hayashi, F.Iraha, N.Shinya, N.Matsuda, D.Kiga, A.Urushibata, M.Shirouzu, K.Oki, T.Kigawa, and S.Yokoyama (2006).
Translation of 'rare' codons in a cell-free protein synthesis system from Escherichia coli.
  J Struct Funct Genomics, 7, 31-36.  
17146616 T.Matsuda, T.Kigawa, S.Koshiba, M.Inoue, M.Aoki, K.Yamasaki, M.Seki, K.Shinozaki, and S.Yokoyama (2006).
Cell-free synthesis of zinc-binding proteins.
  J Struct Funct Genomics, 7, 93.  
15734558 F.Katzen, G.Chang, and W.Kudlicki (2005).
The past, present and future of cell-free protein synthesis.
  Trends Biotechnol, 23, 150-156.  
15830344 K.A.Calhoun, and J.R.Swartz (2005).
Energizing cell-free protein synthesis with glucose metabolism.
  Biotechnol Bioeng, 90, 606-613.  
15991235 K.A.Underwood, J.R.Swartz, and J.D.Puglisi (2005).
Quantitative polysome analysis identifies limitations in bacterial cell-free protein synthesis.
  Biotechnol Bioeng, 91, 425-435.  
15955073 K.Ozawa, M.J.Headlam, D.Mouradov, S.J.Watt, J.L.Beck, K.J.Rodgers, R.T.Dean, T.Huber, G.Otting, and N.E.Dixon (2005).
Translational incorporation of L-3,4-dihydroxyphenylalanine into proteins.
  FEBS J, 272, 3162-3171.  
15741337 N.Handa, T.Terada, Y.Doi-Katayama, H.Hirota, J.R.Tame, S.Y.Park, S.Kuramitsu, M.Shirouzu, and S.Yokoyama (2005).
Crystal structure of a novel polyisoprenoid-binding protein from Thermus thermophilus HB8.
  Protein Sci, 14, 1004-1010.
PDB code: 1wub
15450748 A.S.Spirin (2004).
High-throughput cell-free systems for synthesis of functionally active proteins.
  Trends Biotechnol, 22, 538-545.  
15479237 K.Ozawa, M.J.Headlam, P.M.Schaeffer, B.R.Henderson, N.E.Dixon, and G.Otting (2004).
Optimization of an Escherichia coli system for cell-free synthesis of selectively N-labelled proteins for rapid analysis by NMR spectroscopy.
  Eur J Biochem, 271, 4084-4093.  
15007837 M.C.Jewett, and J.R.Swartz (2004).
Mimicking the Escherichia coli cytoplasmic environment activates long-lived and efficient cell-free protein synthesis.
  Biotechnol Bioeng, 86, 19-26.  
14763830 M.C.Jewett, and J.R.Swartz (2004).
Rapid expression and purification of 100 nmol quantities of active protein using cell-free protein synthesis.
  Biotechnol Prog, 20, 102-109.  
15103643 T.Wada, M.Shirouzu, T.Terada, Y.Kamewari, S.Y.Park, J.R.Tame, S.Kuramitsu, and S.Yokoyama (2004).
Crystal structure of the conserved hypothetical protein TT1380 from Thermus thermophilus HB8.
  Proteins, 55, 778-780.
PDB code: 1iuj
14579367 A.Seto, M.Shirouzu, T.Terada, K.Murayama, S.Kuramitsu, and S.Yokoyama (2003).
Crystal structure of a hypothetical protein, TT1725, from Thermus thermophilus HB8 at 1.7 A resolution.
  Proteins, 53, 768-771.
PDB code: 1j27
12547425 S.Yokoyama (2003).
Protein expression systems for structural genomics and proteomics.
  Curr Opin Chem Biol, 7, 39-43.  
12547032 S.P.Chambers (2002).
High-throughput protein expression for the post-genomic era.
  Drug Discov Today, 7, 759-765.  
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time. Where a reference describes a PDB structure, the PDB code is shown on the right.