DIFFERENTIAL BINDING OF HMG1, HMG2, AND A SINGLE HMG BOX TO CISPLATIN-DAMAGED DNA

The HMG box domain is a DNA binding domain present in the nonhistone c hromosomal proteins HMG1 and HMG2 and in other proteins involved in th e regulation of gene expression. Previous studies have demonstrated th at HMG1 and HMG2 bind with high affinity to DNA modified with the canc er chemotherapeutic drug cisplatin (CDDP). In this report, we compare the binding of full-length HMG1 and HMG2 and the HMG boxes present in these proteins to that of CDDP-DNA. Complexes between HMG1, HMG2, or H MG Box A + B and CDDP-DNA were stable at greater than or equal to 500 mM salt, while complexes between a single HMG box and CDDP-DNA exhibit ed decreased stability, Analysis of a series of HMG1 Box A mutant cons tructs revealed different affinities for CDDP-DNA. Two constructs cont aining a Phe to Ala substitution at position 19 and a Tyr to Gly subst itution at position 71, are noteworthy; these peptides exhibited reduc ed affinity for CDDP-DNA. We have generated a structure of HMG1 Box A and used it, along with the results of our binding studies, to model i ts interaction with CDDP-DNA. HMG1 Box A binds in the minor groove of CDDP-DNA, in agreement with earlier studies. Our model predicts that T yr71 partially intercalates and forms an H bond with the sugar-phospha te backbone. The model also suggests that Phe 19 does not directly int eract with DNA, and hence an Ala substitution at position 19 may alter protein structure. This model should provide a framework for future s tudies examining HMG Box-DNA interactions. (C) 1996 Academic Press, In c.