Co-crystallization of an ETS domain (PU.1) in complex with DNA. Engineering the length of both protein and oligonucleotide.
The PU.1 transcription factor is a member of the ets gene family of regulatory proteins. These molecules play a role in normal development and also have been implicated in malignant processes such as the development of erythroid leukemia. The Ets proteins share a conserved DNA-binding domain (the ETS domain) that recognizes a purine-rich sequence with the core sequence: 5'-C/AGGAA/T-3'. This domain binds to DNA as a monomer, unlike many other DNA-binding proteins. The ETS domain of the PU.1 transcription factor has been crystallized in complex with a 16-base pair oligonucleotide that contains the recognition sequence. The crystals formed in the space group C2 with a = 89.1, b = 101.9, c = 55.6 A, and beta = 111.2 degrees and diffract to at least 2.3 A. There are two complexes in the asymmetric unit. Production of large usable crystals was dependent on the length of both protein and DNA components, the use of oligonucleotides with unpaired A and T bases at the termini, and the presence of polyethylene glycol and zinc acetate in the crystallization solutions. This is the first ETS domain to be crystallized, and the strategy used to crystallize this complex may be useful for other members of the ets family.
Pio
F
F
Cancer Research Center, La Jolla Cancer Research Foundation, California 92037, USA.
Ni
C Z
CZ
Mitchell
R S
RS
Knight
J
J
McKercher
S
S
Klemsz
M
M
Lombardo
A
A
Maki
R A
RA
Ely
K R
KR
D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D017124:Conserved Sequence; D003460:Crystallization; D018360:Crystallography, X-Ray; D004247:DNA / Q000138:chemical synthesis / Q000737:chemistry* / Q000302:isolation & purification; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000737:chemistry* / Q000302:isolation & purification; D011485:Protein Binding; D015202:Protein Engineering; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D015682:Retroviridae Proteins, Oncogenic; D014157:Transcription Factors / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification
D004268:DNA-Binding Proteins; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D015682:Retroviridae Proteins, Oncogenic; D014157:Transcription Factors; C062022:v-Spi-1 protein, Friend spleen focus-forming virus; D004247:DNA