FIGURE 5.

Effect of gp2.5 on leading strand synthesis catalyzed by gp4 and gp5/trx. A, leading strand synthesis catalyzed by wild-type (wt) gp5/trx and the variants with alterations in loops A and B was measured using circular M13 double-stranded DNA bearing a preformed replication fork as shown in the inset. The reactions (10 μl) contained 20 nm DNA; 0.5 mm dATP, dGTP, dCTP, and [α-³²P]dATP; 5 nm gp4 (hexamer); 5 nm of the indicated gp5; 500 nm trx; and 4 μm of gp2.5. After incubation at 37 °C for 10 min, the amount of [α-³²P]dAMP incorporated into DNA was measured and presented as a bar graph. B, gel analysis of products of leading strand DNA synthesis. The radioactive reaction products from A were denatured and analyzed on a 0.6% alkaline-agarose gel by autoradiography.