DNA-BINDING STUDIES OF XSPTSPSZ, DERIVATIVES OF THE INTERCALATING HEPTAD REPEAT OF RNA-POLYMERASE-II

The synthesis, solution conformation, and interaction with DNA of thre e 8-residue peptides structurally related to the heptad repent unit fo und at the C-terminus of RNA polymerase II are reported. Peptides QQ, XQ, and Pe are derived from the parent sequence YSPTSPSY (peptide YY), which was reported to bind to DNA by bisintercalation [M. Suzuki (199 0) Nature, Vol. 344, pp. 562-565], and contain either a 2-quinolyl (Q) , 2-quinoxolyl (X), or 5-phenanthrolyl (P) group in place of the aroma tic side chains of the N- and C-terminal tyrosine residues present in the parent sequence. The combined results of linear dichroism and indu ced CD measurements of peptides QQ, Xe, and Pe with calf thymus DNA ar e consistent with weak binding of the peptides to DNA in a preferred o rientation in which the chromophores are intercalated. Small increases in the melting temperatures of poly[d(A-T)(2)] are also consistent wi th the peptides interacting with DNA. While enzymatic footprinting wit h DNase I showed no protection from cleavage by the enzyme, chemical f ootprinting with fotemustine showed that the peptides modify the react ivity of the major groove, presumably via minor groove binding. Peptid e QQ inhibited fotemustine alkylation significantly more than either X e or Pe, and slightly more than YY. In aqueous solution, nmr experimen ts on QQ, XQ, and PQ show a significant population of conformation in which Ser2-Pro3-Thr4-Ser5 form both type I and type II beta-turn confo rmations in equilibrium with open chain conformations. Nuclear magneti c resonance titration experiments of PQ with (GCGTACGC)(2) showed smal l changes in chemical shifts, consistent with the formation of a weak nonspecific complex. Analogous experiments, using peptides QQ and XQ w ith (GCGTACGC)(2), and peptide YY with (CGTACG)(2), showed no evidence for the interaction of the peptides with these oligonucleotides. Thes e results show that peptides of general structure XSPTSPSZ are weak no nspecific DNA binders that differ significantly from previously charac terized S(T)PXX DNA-binding motifs that are generally AT-selective min or groove binders. (C) 1997 John Wiley & Sons, Inc.