PDBsum entry: 1a8m

Lymphokine

PDB id: 1a8m

Contents

Protein chains

152 a.a. *

Waters ×69

* Residue conservation analysis

PDB id:

1a8m

Name:

Lymphokine

Title:

Tumor necrosis factor alpha, r31d mutant

Structure:

Tumor necrosis factor alpha. Chain: a, b, c. Synonym: tnf-alpha. Engineered: yes. Mutation: yes

Source:

Homo sapiens. Human. Organism_taxid: 9606. Expressed in: escherichia coli. Expression_system_taxid: 562

Biol. unit:

Trimer (from PDB file)

Resolution:

2.30Å R-factor: 0.218 R-free: 0.240

Authors:

C.Reed,Z.-Q.Fu,J.Wu,Y.-N.Xue,R.W.Harrison,M.-J.Chen, I.T.Weber

Key ref:

C.Reed et al. (1997). Crystal structure of TNF-alpha mutant R31D with greater affinity for receptor R1 compared with R2. Protein Eng, 10, 1101-1107. PubMed id: 9488135 DOI: 10.1093/protein/10.10.1101

Date:

27-Mar-98 Release date: 17-Jun-98

Protein chains

P01375  (TNFA_HUMAN) -  Tumor necrosis factor

Seq: 233 a.a.

Struc: 152 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

 Gene Ontology (GO) functional annotation 

• Cellular component: membrane (1 term)
• Biological process: immune response (1 term)
• Biochemical function: tumor necrosis factor receptor binding (1 term)

DOI no: 10.1093/protein/10.10.1101

Protein Eng 10:1101-1107 (1997)

PubMed id: 9488135

Crystal structure of TNF-alpha mutant R31D with greater affinity for receptor R1 compared with R2.

C.Reed, Z.Q.Fu, J.Wu, Y.N.Xue, R.W.Harrison, M.J.Chen, I.T.Weber.

ABSTRACT

Crystal structures have been determined of recombinant human tumor necrosis factor-alpha (TNF-alpha) and its R31D mutant that preferentially binds to TNF receptor R1 with more than seven times the relative affinity of binding to receptor R2. Crystals of the wild-type TNF were of space group P4(1)2(1)2 and had unit cell dimensions of a = b = 94.7 and c = 117.4 A. Refinement of the structure gave an R-factor of 22.3% at 2.5 A resolution. The crystals of TNF R31D mutant diffracted to 2.3 A resolution, and were of identical space group to the wild type with unit cell dimensions of a = b = 95.4 and c = 116.2 A, and the structure was refined to an R-factor of 21.8%. The trimer structures of the wild-type and mutant TNF were similar with a root mean square (r.m.s.) deviation of 0.56 A for Calpha atoms; however, the subunits within each trimer were more variable with an average r.m.s. deviation of 1.00 A on Calpha atoms for pairwise comparison of subunits. Model complexes of TNF with receptors R1 and R2 have been used to predict TNF-receptor interactions. Arg31 in all three subunits of wild-type TNF is predicted to form an ionic interaction with the equivalent glutamic acid in both receptors R1 and R2. Asp31 of the TNF R31D mutant is predicted to interact differently with the two receptors. The side chain of Asp31 in two subunits of the TNF mutant is predicted to form hydrogen bond interactions with Ser59 or Cys70 of R1, while it has no predicted interactions with R2. The loss of three strong ionic interactions of Arg31 and the electrostatic repulsion of Asp31 with Glu in the receptors is consistent with the reduced binding of the R31D mutant to both receptors relative to wild-type TNF. The replacement of these ionic interactions by two weaker hydrogen bond interactions between Asp31 of the R31D mutant and R1, compared with no interactions with R2, is in agreement with the observed preferential binding of the R31D mutant to R1 over R2. Analysis of the structure and function of receptor-discriminating mutants of TNF will help understand the biological role of TNF and facilitate its use as an antitumor agent.