Cs. Chow et al., A SINGLE HMG DOMAIN IN HIGH-MOBILITY GROUP-1 PROTEIN BINDS TO DNAS ASSMALL AS 20 BASE-PAIRS CONTAINING THE MAJOR CISPLATIN ADDUCT, Biochemistry, 34(9), 1995, pp. 2956-2964
Proteins containing a relatively new DNA-binding motif known as the hi
gh-mobility group (HMG) domain bind specifically to DNA modified by th
e anticancer drug cisplatin, but not to unmodified DNA (McA'Nulty & Li
ppard, 1995). Southwestern-blot analyses of the binding of proteolytic
fragments of HMG1 to a 123-bp globally platinated DNA demonstrate tha
t the HMG domains A and B of HMG1 are responsible for its specific int
eractions with cisplatin-modified DNA. An 81 amino acid recombinant pr
otein representing a single HMG motif, HMG1 domain B, binds with an af
finity (Kd = 10(-7) M) equal to that of HMG1 itself to 92- and 100-bp
DNAs containing the major adduct of cisplatin, a cis-[Pt(NH3)(2)-{d(Gp
G)-N7(1), -N7(2)}] intrastrand cross-link, at a specific site. The iso
lated HMG domain B binds with comparable affinity to cisplatin-modifie
d DNAs having as few as 20 bp. The related human mitochondrial HMG dom
ain protein mtTFA also recognizes the 123-bp globally platinated DNA,
providing further evidence that HMG domains are responsible for modula
ting binding of this class of proteins to cisplatin-modified DNA. This
work provides direct biochemical evidence in support of conclusions d
rawn previously from analyses of sequence conservation (Bruhn et al.,
1992) that HMG domains are the key elements in protein binding to cisp
latin-modified DNA.