STRUCTURE OF TAQ POLYMERASE WITH DNA AT THE POLYMERASE ACTIVE-SITE

THE DNA polymerase from Thermus aquaticus (Tag polymerase) is homologo us to Escherichia coli DNA polymerase I (Pol I) and likewise has domai ns responsible for DNA polymerase and 5' nuclease activities(1,2). The structures of the polymerase domains of Tag polymerase and of the Kle now fragment (KF) of Pol I are almost identical, whereas the structure of a vestigial editing 3'-5' exonuclease domain of Tag polymerase tha t lies between the other two domains is dramatically altered, resultin g in the absence of this activity in the thermostable enzyme(2). The s tructures have been solved for editing complexes between KF and single -stranded DNA(3,4) and for duplex DNA with a 3' overhanging single str and(5), but not for a complex containing duplex DNA at the polymerase active-site, Here we present the co-crystal structure of Taq polymeras e with a blunt-ended duplex DNA bound to the polymerase active-site cl eft; the DNA neither bends nor goes through the large polymerase cleft , and the structural form of the bound DNA is between the B and A form s. A wide minor groove allows access to protein side chains that hydro gen-bond to the N3 of purines and the O2 of pyrimidines at the blunt-e nd terminus. Part of the DNA bound to the polymerase site shares a com mon binding site with DNA bound to the exonuclease site, but they are translated relative to each other by several angstroms along their hel ix axes.