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PDBsum entry 2e7a

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protein Protein-protein interface(s) links
Cytokine PDB id
2e7a
Jmol
Contents
Protein chains
150 a.a. *
142 a.a. *
Waters ×237
* Residue conservation analysis
PDB id:
2e7a
Name: Cytokine
Title: Tnf receptor subtype one-selective tnf mutant with antagonistic activity
Structure: Tumor necrosis factor. Chain: a, b, c. Synonym: tnf. Engineered: yes. Mutation: yes
Source: Homo sapiens. Human. Organism_taxid: 9606. Expressed in: escherichia coli bl21(de3). Expression_system_taxid: 469008. Other_details: this mutant was created by phage display system
Resolution:
1.80Å     R-factor:   0.198     R-free:   0.239
Authors: Y.Mukai,Y.Yamagata,Y.Tsutsumi
Key ref:
H.Shibata et al. (2008). Creation and X-ray structure analysis of the tumor necrosis factor receptor-1-selective mutant of a tumor necrosis factor-alpha antagonist. J Biol Chem, 283, 998. PubMed id: 18003610 DOI: 10.1074/jbc.M707933200
Date:
09-Jan-07     Release date:   13-Nov-07    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chain
Pfam   ArchSchema ?
P01375  (TNFA_HUMAN) -  Tumor necrosis factor
Seq:
Struc:
233 a.a.
150 a.a.*
Protein chains
Pfam   ArchSchema ?
P01375  (TNFA_HUMAN) -  Tumor necrosis factor
Seq:
Struc:
233 a.a.
142 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 24 residue positions (black crosses)

 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     membrane   1 term 
  Biological process     immune response   1 term 
  Biochemical function     tumor necrosis factor receptor binding     1 term  

 

 
DOI no: 10.1074/jbc.M707933200 J Biol Chem 283:998 (2008)
PubMed id: 18003610  
 
 
Creation and X-ray structure analysis of the tumor necrosis factor receptor-1-selective mutant of a tumor necrosis factor-alpha antagonist.
H.Shibata, Y.Yoshioka, A.Ohkawa, K.Minowa, Y.Mukai, Y.Abe, M.Taniai, T.Nomura, H.Kayamuro, H.Nabeshi, T.Sugita, S.Imai, K.Nagano, T.Yoshikawa, T.Fujita, S.Nakagawa, A.Yamamoto, T.Ohta, T.Hayakawa, T.Mayumi, P.Vandenabeele, B.B.Aggarwal, T.Nakamura, Y.Yamagata, S.Tsunoda, H.Kamada, Y.Tsutsumi.
 
  ABSTRACT  
 
Tumor necrosis factor-alpha (TNF) induces inflammatory response predominantly through the TNF receptor-1 (TNFR1). Thus, blocking the binding of TNF to TNFR1 is an important strategy for the treatment of many inflammatory diseases, such as hepatitis and rheumatoid arthritis. In this study, we identified a TNFR1-selective antagonistic mutant TNF from a phage library displaying structural human TNF variants in which each one of the six amino acid residues at the receptor-binding site (amino acids at positions 84-89) was replaced with other amino acids. Consequently, a TNFR1-selective antagonistic mutant TNF (R1antTNF), containing mutations A84S, V85T, S86T, Y87H, Q88N, and T89Q, was isolated from the library. The R1antTNF did not activate TNFR1-mediated responses, although its affinity for the TNFR1 was almost similar to that of the human wild-type TNF (wtTNF). Additionally, the R1antTNF neutralized the TNFR1-mediated bioactivity of wtTNF without influencing its TNFR2-mediated bioactivity and inhibited hepatic injury in an experimental hepatitis model. To understand the mechanism underlying the antagonistic activity of R1antTNF, we analyzed this mutant using the surface plasmon resonance spectroscopy and x-ray crystallography. Kinetic association/dissociation parameters of the R1antTNF were higher than those of the wtTNF, indicating very fast bond dissociation. Furthermore, x-ray crystallographic analysis of R1antTNF suggested that the mutation Y87H changed the binding mode from the hydrophobic to the electrostatic interaction, which may be one of the reasons why R1antTNF behaved as an antagonist. Our studies demonstrate the feasibility of generating TNF receptor subtype-specific antagonist by extensive substitution of amino acids of the wild-type ligand protein.
 
  Selected figure(s)  
 
Figure 6.
FIGURE 6. Overall structures of R1antTNF and wtTNF. A, refined structure of the R1antTNF trimer (green). Blue spheres show the mutated residues(amino acids 84-89) in R1antTNF. This structure is registered in the PDB (PDB code 2E7A). B, structure of the wtTNF trimer (gray). This structure has been published, and its PDB code is 1TNF. C, model structures of the TNF-TNFR1 complexes. Each TNF is superposed on the LT- derived from the LT- -TNFR1 complex (PDB code 1TNR). TNF binds to three R1 monomers on the cell surface. TNFR1s are shown using red schematics. Superposition of the structures of the wtTNF and R1antTNF was performed using the ccp4i program.
Figure 7.
FIGURE 7. Structural difference between the receptor binding region of the R1antTNF and wtTNF. A, interaction between the wtTNF (gray) and TNFR1 (red). White layer depicts the molecular surface of the TNFR1. Hydrophobic interaction is formed between the Tyr-87 and molecular pocket in the TNFR1 (Leu-67, Leu-71, Ala-62, and Ser-63). B, interaction between the R1antTNF (green) and TNFR1 (red). Yellow broken lines show the possible interactions of the R1antTNF His-87 with the receptor Ser-63 and Glu-64. In this simulation, the side chains of each structure were rotated to fit the predicted interaction. Stable structures of these rotamers were constructed using the O program.
 
  The above figures are reprinted by permission from the ASBMB: J Biol Chem (2008, 283, 998) copyright 2008.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
20036293 T.Nomura, Y.Abe, H.Kamada, H.Shibata, H.Kayamuro, M.Inoue, T.Kawara, S.Arita, T.Furuya, T.Yamashita, K.Nagano, T.Yoshikawa, Y.Yoshioka, Y.Mukai, S.Nakagawa, M.Taniai, T.Ohta, S.Serada, T.Naka, S.Tsunoda, and Y.Tsutsumi (2011).
Therapeutic effect of PEGylated TNFR1-selective antagonistic mutant TNF in experimental autoimmune encephalomyelitis mice.
  J Control Release, 149, 8.  
20111721 J.Dong, Y.Gao, Y.Liu, J.Shi, J.Feng, Z.Li, H.Pan, Y.Xue, C.Liu, B.Shen, N.Shao, and G.Yang (2010).
The protective antibodies induced by a novel epitope of human TNF-alpha could suppress the development of collagen-induced arthritis.
  PLoS One, 5, e8920.  
20371990 K.Nagano, S.Imai, S.Nakagawa, S.Tsunoda, and Y.Tsutsumi (2010).
[From disease proteomics to biomarker development-establishment of antibody proteomics technology and exploration of cancer-related biomarkers].
  Yakugaku Zasshi, 130, 487-492.  
21314616 L.N.Shingarova, E.F.Boldyreva, S.A.Yakimov, S.V.Guryanova, D.A.Dolgikh, S.A.Nedospasov, and M.P.Kirpichnikov (2010).
Novel mutants of human tumor necrosis factor with dominant-negative properties.
  Biochemistry (Mosc), 75, 1458-1463.  
20017116 R.L.Rich, and D.G.Myszka (2010).
Grading the commercial optical biosensor literature-Class of 2008: 'The Mighty Binders'.
  J Mol Recognit, 23, 1.  
20117833 T.Morishige, Y.Yoshioka, H.Inakura, A.Tanabe, X.Yao, S.Tsunoda, Y.Tsutsumi, Y.Mukai, N.Okada, and S.Nakagawa (2010).
Creation of a LIGHT mutant with the capacity to evade the decoy receptor for cancer therapy.
  Biomaterials, 31, 3357-3363.  
20046067 T.Nomura, Y.Abe, Y.Yoshioka, S.Nakagawa, S.Tsunoda, and Y.Tsutsumi (2010).
[Creation of TNFR1-selective antagonist and its therapeutic effects]
  Yakugaku Zasshi, 130, 63-68.  
19853911 Y.Yoshioka, H.Watanabe, T.Morishige, X.Yao, S.Ikemizu, C.Nagao, S.Ahmad, K.Mizuguchi, S.Tsunoda, Y.Tsutsumi, Y.Mukai, N.Okada, and S.Nakagawa (2010).
Creation of lysine-deficient mutant lymphotoxin-alpha with receptor selectivity by using a phage display system.
  Biomaterials, 31, 1935-1943.  
19646748 H.Kayamuro, Y.Abe, Y.Yoshioka, K.Katayama, T.Nomura, T.Yoshida, K.Yamashita, T.Yoshikawa, Y.Kawai, T.Mayumi, T.Hiroi, N.Itoh, K.Nagano, H.Kamada, S.Tsunoda, and Y.Tsutsumi (2009).
The use of a mutant TNF-alpha as a vaccine adjuvant for the induction of mucosal immune responses.
  Biomaterials, 30, 5869-5876.  
19765818 H.Shibata, Y.Yoshioka, Y.Abe, A.Ohkawa, T.Nomura, K.Minowa, Y.Mukai, S.Nakagawa, M.Taniai, T.Ohta, H.Kamada, S.Tsunoda, and Y.Tsutsumi (2009).
The treatment of established murine collagen-induced arthritis with a TNFR1-selective antagonistic mutant TNF.
  Biomaterials, 30, 6638-6647.  
19652499 Y.Abe (2009).
[Development of novel DDS technologies for optimized protein therapy by creating functional mutant proteins with antagonistic activity]
  Yakugaku Zasshi, 129, 933-939.  
19386778 Y.Mukai, T.Nakamura, Y.Yoshioka, H.Shibata, Y.Abe, T.Nomura, M.Taniai, T.Ohta, S.Nakagawa, S.Tsunoda, H.Kamada, Y.Yamagata, and Y.Tsutsumi (2009).
Fast binding kinetics and conserved 3D structure underlie the antagonistic activity of mutant TNF: useful information for designing artificial proteo-antagonists.
  J Biochem, 146, 167-172.
PDB code: 2zpx
  19255488 Y.Mukai, T.Nakamura, Y.Yoshioka, S.Tsunoda, H.Kamada, S.Nakagawa, Y.Yamagata, and Y.Tsutsumi (2009).
Crystallization and preliminary X-ray analysis of the tumour necrosis factor alpha-tumour necrosis factor receptor type 2 complex.
  Acta Crystallogr Sect F Struct Biol Cryst Commun, 65, 295-298.  
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.