Purification, crystallization and preliminary x-ray crystallographic studies of RAIDD Death-Domain (DD)

Caspase-2 activation by formation of PIDDosome is critical for genotoxic stress induced apoptosis. PIDDosome is composed of three proteins, RAIDD, PIDD, and Caspase-2. RAIDD is an adaptor protein containing an N-terminal Caspase-Recruiting-Domain (CARD) and a C-terminal Death-Domain (DD). Its intera...

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Bibliographic Details
Published in:International journal of molecular sciences 2009-06, Vol.10 (6), p.2501-2509
Main Authors: Jang, Tae-ho, Park, Hyun Ho
Format: Article
Language:English
Subjects:
Apoptosis
Chromatography
Chromatography, Affinity
Cloning
Cloning, Molecular
CRADD Signaling Adaptor Protein - chemistry
CRADD Signaling Adaptor Protein - genetics
CRADD Signaling Adaptor Protein - isolation & purification
Crystallization
Crystallography, X-Ray
Crystals
Death-Domain (DD)
E coli
Humans
Kinases
Molecular structure
Physiology
PIDDosome
Protein Structure, Tertiary
Proteins
RAIDD
Recombinant Proteins - biosynthesis
Recombinant Proteins - chemistry
Recombinant Proteins - isolation & purification
Selenium
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Summary:Caspase-2 activation by formation of PIDDosome is critical for genotoxic stress induced apoptosis. PIDDosome is composed of three proteins, RAIDD, PIDD, and Caspase-2. RAIDD is an adaptor protein containing an N-terminal Caspase-Recruiting-Domain (CARD) and a C-terminal Death-Domain (DD). Its interactions with Caspase-2 and PIDD through CARD and DD respectively and formation of PIDDosome are important for the activation of Caspase-2. RAIDD DD cloned into pET26b vector was expressed in E. coli cells and purified by nickel affinity chromatography and gel filtration. Although it has been known that the most DDs are not soluble in physiological condition, RAIDD DD was soluble and interacts tightly with PIDD DD in physiological condition. The purified RAIDD DD alone has been crystallized. Crystals are trigonal and belong to space group P3(1)21 (or its enantiomorph P3(2)21) with unit-cell parameters a = 56.3, b = 56.3, c = 64.9 A and gamma = 120 degrees . The crystals were obtained at room temperature and diffracted to 2.0 A resolution.
ISSN:1422-0067
1661-6596
1422-0067
DOI:10.3390/ijms10062501