Purification, crystallization and preliminary X-ray diffraction analysis of the HMG domain of Sox17 in complex with DNA.
Sox17 is a member of the SRY-related high-mobility group (HMG) of transcription factors that have been shown to direct endodermal differentiation in early mammalian development. The LAMA1 gene encoding the alpha-chain of laminin-1 has been reported to be directly bound and regulated by Sox17. This paper describes the details of initial crystallization attempts with the HMG domain of mouse Sox17 (mSox17-HMG) with a 16-mer DNA element derived from the LAMA1 enhancer and optimization strategies to obtain a better diffracting crystal. The best diffracting crystal was obtained in a condition containing 0.1 M Tris-HCl pH 7.4, 0.2 M MgCl(2), 30% PEG 3350 using the hanging-drop vapour-diffusion method. A highly redundant in-house data set was collected to 2.75 A resolution with 99% completeness. The presence of the mSox17-HMG-DNA complex within the crystals was confirmed and Matthews analysis indicated the presence of one complex per asymmetric unit.
Ng
Calista Keow Leng
CK
Laboratory for Structural Biochemistry, Genome Institute of Singapore, Genome, 60 Biopolis Street, Singapore 138672, Singapore.
Palasingam
Paaventhan
P
Venkatachalam
Rajakannan
R
Baburajendran
Nithya
N
Cheng
Jason
J
Jauch
Ralf
R
Kolatkar
Prasanna R
PR
D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D024002:HMGB Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D055760:SOXF Transcription Factors / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D014961:X-Ray Diffraction
D024002:HMGB Proteins; D055760:SOXF Transcription Factors; C511606:Sox17 protein, mouse; D004247:DNA