Human chromodomain protein, Y chromosome, 2 related

Target ID:

CDY

Aliases:

CDYCDY2MGC163298MGC163300

Deposition Date:

Tuesday 31st January 2006

Families:

Transferases

Related Diseases:

Chromatin and epigenetics

Structure type:

apo

Download Coordinates:

2FW2

Authors:

J.R.MIN,T.ANTOSHENKO,H.WU,J.WEIGELT,M.SUNDSTROM,C.ARROWSMITH,A.M.EDWARDS,A.BOCHKAREV,A.N.PLOTNIKOV,STRUCTURAL GENOMICS CONSORTIUM (SGC)

Site:

Toronto

2FW2

tabs

Structure Details

Histone acetyltransferases play an important role in the signal transduction, chromatin remodeling as well as DNA replication, transcription and repair.
The CDY family of genes is of special interest because some of them are included in chromosome Y microdeletions detected among infertile men and are apparently involved in the spermiogenesis.
There are six genes (CDY, CDY1, CDY1B, CDY2, CDYL and CDYL2) in the human genome.
It has been shown that the progenitor of the CDY gene family arose de novo in the mammalian ancestor via domain accretion.
This progenitor later duplicated to generate CDYL and CDYL2, two autosomal genes found in all extant mammals. Prior to human-mouse divergence, a processed CDYL transcript retroposed onto the Y chromosome to create CDY.
The CDY genes encode the proteins containing two functional domains - a chromodomain, a sequence motif involved in chromatin binding, and a catalytic domain found in many CoA-dependent acetyltransferases.
Here we have solved the crystal structure of the catalytic domain of human CDY protein at 2.2 Å resolution.

Materials and Methods

CDY

PDB:2FW2

Revision

Revision Type:created
Revised by:created
Revision Date:created
Entry Clone Accession:GI:47271406
Entry Clone Source:MGC
SGC Clone Accession:
Tag:N-terminal: His-tag with integrated thrombin protease site: MGSSHHHHHHSSGLVPRGS
Host:E.coli BL21 (DE3) codon plus RIL (Stratagen).

Construct

Prelude:
Sequence:ITYRDIVVKKEDGFTQIVLSTRSTEKNALNTEVIKEMVNALNSAAADDSKLVLFSAAGSVFCCGLDFGYFVRHLRNDRNTASLEMVDTIKNFVNTFIQFKKPIVVSVNGPAIGLGASILPLCDLVWANEKAWFQTPYTTFGQSPDGCSSITFPKMMGKASANEMLIAGRKLTAREACAKGLVSQVFLTGTFTQEVMIQIKELASYNAIVLEECKALVRCNIKLELEQANERECEVLRKIWSSAQGIESMLKYVENKIDEF
Vector:pET28a-LIC

Growth

Medium:
Antibiotics:
Procedure:CDY was expressed in E.coli BL21 (DE3) codon plus RIL in TB medium in the presence of 50 µg/mL of kanamycin. Cell were grown at 37oC to an OD600 of 0.8 and induced by isopropyl-1-thio-D-galactopyranoside (IPTG), final concentration 1 mM and incubated overnight at 15oC.

Purification

Procedure

Extraction

Procedure
Cells were harvested by centrifugation at 7,000 rpm. The cell pellets were frozen in liquid nitrogen and stored at -80°C. For the purification the cell paste was thawed and resuspended in lysis buffer (50 mM Hepes, pH 7.5, 0.5 M NaCl, 5 mM imidazol, 2 mM ß-mercaptoethanol, 5% glycerol) with protease inhibitor (0.1mM phenylmethyl sulfonyl fluoride, PMSF). The cells were lysed by passing through Microfluidizer (Microfluidics Corp.) at 20,000 psi.
Concentration:
Ligand
MassSpec:
Crystallization:Purified CDY was crystallized using sitting drop vapor diffusion method drop at 20 °C by mixing 1.0 µL of the protein solution with 1.0 µL of the reservoir solution containing 8% PEG 4000, 0.1M Na Acetate, 0.1M Na Acetate pH 4.6.
NMR Spectroscopy:
Data Collection:
Data Processing: