PDBsum entry 2ocf

Hormone/growth factor

PDB id: 2ocf

Contents

Protein chains

233 a.a. *

93 a.a. *

Ligands

EST

Waters ×11

* Residue conservation analysis

PDB id:

2ocf

Name:

Hormone/growth factor

Title:

Human estrogen receptor alpha ligand-binding domain in complex with estradiol and the e2#23 fn3 monobody

Structure:

Estrogen receptor. Chain: a. Fragment: ligand binding domain, residues 298-595. Synonym: er, estradiol receptor, er-alpha. Engineered: yes. Mutation: yes. Fibronectin. Chain: d. Fragment: e2#23 fn3 monobody, fibronectin type iii domain 10,

Source:

Homo sapiens. Human. Organism_taxid: 9606. Gene: esr1, esr, nr3a1. Expressed in: escherichia coli bl21(de3). Expression_system_taxid: 469008. Gene: cig, fn1, fn. Expressed in: escherichia coli. Expression_system_taxid: 562

Resolution:

2.95Å R-factor: 0.194 R-free: 0.251

Authors:

S.S.Rajan,S.M.Kuruvilla,S.K.Sharma,Y.Kim,J.Huang,A.Koide,S.Koide, A.Joachimiak,G.L.Greene

Key ref:

A.Koide et al. (2002). Probing protein conformational changes in living cells by using designer binding proteins: application to the estrogen receptor. Proc Natl Acad Sci U S A, 99, 1253-1258. PubMed id: 11818562

Date:

20-Dec-06 Release date: 06-Nov-07

Protein chain

P03372  (ESR1_HUMAN) -  Estrogen receptor from Homo sapiens

Seq: 595 a.a.

Struc: 233 a.a.*

Protein chain

P02751  (FINC_HUMAN) -  Fibronectin from Homo sapiens

Seq: 2477 a.a.

Struc: 93 a.a.*

* PDB and UniProt seqs differ at 11 residue positions (black crosses)

Proc Natl Acad Sci U S A 99:1253-1258 (2002)

PubMed id: 11818562

Probing protein conformational changes in living cells by using designer binding proteins: application to the estrogen receptor.

A.Koide, S.Abbatiello, L.Rothgery, S.Koide.

ABSTRACT

A challenge in understanding the mechanism of protein function in biology is to establish the correlation between functional form in the intracellular environment and high-resolution structures obtained with in vitro techniques. Here we present a strategy to probe conformational changes of proteins inside cells. Our method involves: (i) engineering binding proteins to different conformations of a target protein, and (ii) using them to sense changes in the surface property of the target in cells. We probed ligand-induced conformational changes of the estrogen receptor alpha (ER alpha) ligand-binding domain (LBD). By using yeast two-hybrid techniques, we first performed combinatorial library screening of "monobodies" (small antibody mimics using the scaffold of a fibronectin type III domain) for clones that bind to ER alpha and then characterized their interactions with ER alpha in the nucleus, the native environment of ER alpha, in the presence of various ligands. A library using a highly flexible loop yielded monobodies that specifically recognize a particular ligand complex of ER alpha, and the pattern of monobody specificity was consistent with the structural differences found in known crystal structures of ER alpha-LBD. A more restrained loop library yielded clones that bind both agonist- and antagonist-bound ER alpha. Furthermore, we found that a deletion of the ER alpha F domain that is C-terminally adjacent to the LBD increased the crossreactivity of monobodies to the apo-ER alpha-LBD, suggesting a dynamic nature of the ER alpha-LBD conformation and a role of the F domain in restraining the LBD in an inactive conformation.