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GrainGenes Reference Report: PMB-46-193

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Reference
PMB-46-193
Title
DNA binding mediated by the wheat HMGa protein: a novel instance of selectivity against alternating GC sequence
Journal
Plant Molecular Biology
Year
2001
Volume
46
Pages
193-204
Author
Leng CY
Hock PK
Kini RM
Abstract
Summary: The high-mobility-group (HMG) chromosomal protein wheat HMGa was purified to homogeneity and tested for its binding characteristics to double-stranded DNA. Wheat HMGa was able to bind to P268, an A/T-rich fragment derived from the pea plastocyanin gene promoter, producing a small mobility shift in gel retardation assays where the bound complex was sensitive to addition of proteinase K but resistant to heat treatment of the protein, consistent with the identity of wheat HMGa as a putative HMG-I/Y protein. Gel retardation assays and south-western hybridization analysis revealed that wheat HMGa could selectively interact with the DNA polynucleotides poly(dA).poly(dT), poly(dAdT).poly(dAdT), and poly(dG).poly(dC), but not with poly(dGdC).poly(dGdC). Surface plasmon resonance analysis determined the kinetic and affinity constants of sensor chip-immobilized wheat HMGa for double-stranded DNA 10-mers, revealing a good affinity of the protein for various dinucleotide combinations, except that of alternating GC sequence. Thus contrary to prior reports of a selectivity of wheat HMGa for A/T-rich DNA, the protein appears to be able to interact with sequences containing guanine and cytosine residues as well, except where G/C residues alternate directly in the primary sequence
Keyword
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a/T-rich
affinities
affinity
alternate
assay
assays
binding characteristics
canavalia-gladiata-dc
complexes
cytosine
dinucleotide
dna
dna binding
g/C-rich
gene
gene promoter
group chromosomal-proteins
heat
heat treatment
high-mobility-group (HMG) proteins
histone h1
hybridization
interact
its
k
kinetic-analysis
minor-groove
mobility
mobility-group proteins
pea
plant science
plastocyanin gene promoter
polynucleotides
positive regulatory region
promoter
protein
proteinase
residue
residues
resistant
selectivity
sensor
sequence
southwestern hybridization
surface
surface plasmon resonance
surface plasmon resonance analysis
transgenic tobacco
wheat HMGa