PDBsum entry 1uzc

Nuclear protein

PDB id: 1uzc

Contents

Protein chain

69 a.a. *

* Residue conservation analysis

PDB id:

1uzc

Name:

Nuclear protein

Title:

The structure of an ff domain from human hypa/fbp11

Structure:

Hypothetical protein flj21157. Chain: a. Fragment: ff domain, residues 250-319. Synonym: huntingtin-interacting protein hypa/fbp11. Engineered: yes

Source:

Homo sapiens. Human. Organism_taxid: 9606. Expressed in: escherichia coli bl21(de3). Expression_system_taxid: 469008. Expression_system_variant: c41.

NMR struc:

23 models

Authors:

M.D.Allen,P.Jemth,A.Friedler,O.Schon,M.Bycroft

Key ref:

M.Allen et al. (2002). The structure of an FF domain from human HYPA/FBP11. J Mol Biol, 323, 411-416. PubMed id: 12381297 DOI: 10.1016/S0022-2836(02)00968-3

Date:

09-Mar-04 Release date: 05-Apr-04

Supersedes:

1h40

Protein chain

O75400  (PR40A_HUMAN) -  Pre-mRNA-processing factor 40 homolog A from Homo sapiens

Seq: 957 a.a.

Struc: 69 a.a.

DOI no: 10.1016/S0022-2836(02)00968-3

J Mol Biol 323:411-416 (2002)

PubMed id: 12381297

The structure of an FF domain from human HYPA/FBP11.

M.Allen, A.Friedler, O.Schon, M.Bycroft.

ABSTRACT

The FF domain is a 60 amino acid residue phosphopeptide-binding module found in a variety of eukaryotic proteins including the transcription elongation factor CA150, the splicing factor Prp40 and p190RHOGAP. We have determined the structure of an FF domain from HYPA/FBP11. The domain is composed of three alpha helices arranged in an orthogonal bundle with a 3(10) helix in the loop between the second and third alpha helices. The structure differs from those of other phosphopeptide-binding domains and represents a novel phosphopeptide-binding fold.

Selected figure(s)

Figure 1.
Figure 1. (a) An overlay of the backbone atoms of the 25 lowest-energy NMR structures. The coding sequence for residues 357-425 of SPTREMBL O75400 corresponding to the FF domain was amplified by PCR from IMAGE cDNA clone 731611 (obtained from the MRC HGMP Resource Centre) by standard methods and cloned into a pRSET-derived pHisGro vector. This was used to over-express a soluble histidine-tagged GroEL apical domain/FF domain fusion protein in Escherichia coli. The fusion protein was purified under native conditions using NTA agarose (Qiagen). After cleavage with thrombin, the FF domain was purified using ion-exchange chromatography and gel-filtration. The samples for NMR spectroscopy typically contained 2.5 mM human FF domain in 90% H[2]O/10% 2H[2]O containing 50 mM KCl, 50 mM potassium phosphate (pH 6.0) at 298 K. The NMR spectra were assigned using standard NMR methods.[18. and 19.] The assignments have been deposited in the BioMagResBank under accession numbers PDB 1H40 BMRDB 5537. A set of distance constraints were derived from a series of NOESY spectra recorded in H[2]O and 2H[2]O with mixing times of 150 ms. The NOESY spectrum was integrated according to the cross-peak strengths and calibrated by comparison with NOE connectivities obtained for standard inter-residue distances within an a helix. After calibration, the NOE constraints were classified into the following categories: strong, medium, weak and very weak, corresponding to inter-proton distance constraints of 1.8-2.8 Å, 1.8-3.5 Å, 1.8-4.75 Å, and 2.5-6.0 Å, respectively. Hydrogen bond constraints were included for a number of backbone NH groups whose signals were observed in a 2D 1H-15N-HSQC recorded in 99.996% 2H[2]O at 298 K (pH 5.0). For hydrogen bond partners, two distance constraints were used where the distance (D)H-O(A) corresponded to 1.5-2.5 Å and (D)N-O(A) to 2.5-3.5 Å. Torsional angle constraints were obtained from an analysis of C', N, C^a Ha and C^b chemical shifts using the program TALOS.[20.] The three-dimensional structure of the FF domain was calculated using a dynamic simulated annealing protocol based upon the work of Nilges et al.[21.] in the program XPLOR (Brünger, A. T. (1992). X-PLOR Version 3.1: a system for cystallography and NMR, Yale University, New Haven, CT). The coordinates have been deposited in the protein structure database, entry. (b) A ribbon representation of the lowest-energy structure prepared using the program MOLSCRIPT.[22.] (c) A ribbon representation of the C-terminal region of human phosphatase 2C alpha prepared using the program MOLSCRIPT. [22.] Note there is a break in the electron density in the loop between the first and second helices.

Figure 2.
Figure 2. (a) Ribbon representation of the FF domain with the side-chains of the structural ensemble for the conserved residues in the hydrophobic core of the domain. (b) Structure-based sequence alignment of selected FF domains. Proteins included are the WW/FF domain-containing proteins that have been shown to bind to RNAP II CTD via their FF domains (mouse HYPA/FBP11(Q9R1C7), Saccharomyces cerevisiae PRP40 (PR40_YEAST), human CA150 (O14776)), human RHOGAP 190-A (Q9NRY4) and human RHOGAP 190-B (Q13017). Residues that are conserved in 50% of FF domains are indicated.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2002, 323, 411-416) copyright 2002.

Figures were selected by an automated process.