Nuclear matrix protein ARBP recognizes a novel DNA sequence motif with high affinity.
ARBP is a nuclear protein that specifically binds to matrix/scaffold attachment regions (MARs/SARs). Here we characterize by DNase I footprinting, dimethyl sulfate protection, and mobility shift assays two binding sites for ARBP within a chicken lysozyme MAR fragment. Our results indicate that ARBP recognizes a novel DNA sequence motif containing the central sequence 5'-GGTGT-3' and flanking AT-rich sequences. Binding occurs through major groove contacts to two guanines of the central sequence. Collective and single-base substitutions in the 5'-GGTGT-3' core motif result in loss or significant reductions of ARBP binding, underscoring the importance of the GC-rich core sequence. Structural elements of the sequence motif are probably also recognized. The affinity of ARBP to both binding sites is surprisingly high [KD = (2-6) x 10(-10) M]. High-affinity recognition of the identified DNA motif in MARs/SARs by ARBP is likely an important feature in the domain organization of chromatin.
Buhrmester
H
H
Institut für Physiologische Chemie, Universitäts-Krankenhaus Eppendorf, Hamburg, FR Germany.
von Kries
J P
JP
Strätling
W H
WH
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002645:Chickens; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004252:DNA Mutational Analysis; D003850:Deoxyribonuclease I; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D007700:Kinetics; D036961:Matrix Attachment Region Binding Proteins; D008969:Molecular Sequence Data; D009113:Muramidase / Q000737:chemistry / Q000378:metabolism; D015530:Nuclear Matrix / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D015183:Restriction Mapping; D012261:Ribonucleoproteins / Q000302:isolation & purification / Q000378:metabolism*; D017384:Sequence Deletion; D013379:Substrate Specificity
C439935:ARBP protein, chicken; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D036961:Matrix Attachment Region Binding Proteins; D010446:Peptide Fragments; D011994:Recombinant Proteins; D012261:Ribonucleoproteins; D004247:DNA; D003850:Deoxyribonuclease I; D009113:Muramidase