Lecture 4 For BGYA01

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Biological Sciences
Clare Hasenkampf

Sept. 20, 2007 Lecture 4 BGYA01 In the last class we were talking about DNA replication. Clicker question An origin of replication a) occurs at the end of the double stranded DNA b) has two RNA primersorigin c) occurs in the interior of the double stranded DNA d) has one RNA primer per origin e) answers b and c are both true We know that DNA replication is initiated at origins of replication (which are in the interior of the double stranded DNA molecule) and that replication proceeds bi-directionally from the origin. Figure 11.13, page 244 and homemade figure all Bidirectional replication creates a growing replication bubble, but the textbook is lazy and only shows events for half of the bubble one replication fork. We also know that the first nucleic acid synthesized at the origin is actually RNA. An RNA primer is made for each fork (ie two RNA primers per origin). homemade figure frame d and Figure 11.16, page 246 Once the RNA primers are laid down at the origin, things go smoothly on one half of each fork. For each fork Figure 11.18, page 247- One newly synthesized strand has the correct polarity to readily continue replication from the original RNA primer; each time a nucleotide is added on, there is a free 3OH group on that nucleotide to add on to. We call such a newly created strand a leading strand. We also say DNA is create 5 to 3 (from the 5 end to the (growing) 3 end. For this terminology the reference is the newly synthesized DNA. As helicase continually opens up the DNA helix of this fork, replication occurs continuously on the leading strands. We say replication on the leading strands is continuous . The other portion of the same fork (the one that uses the antiparallel strand of DNA as the template) has a more difficult time. Why? It is because that region has a template, but no free 3OH group (to add onto) in the direction the fork is opening. ). textbook figure 11.18, page 247 top panel So at the moment no replication can occur there (study guide replication figure, part D Instead replication lags on this side of the fork until the DNA helix gets more opened up. We call the strand that will eventually be made on this portion of the fork, the lagging strand. If you look at the homemade figure you should be able to see that there are actually two leading and two lagging strands per origin of replication. www.notesolution.comSo, how does the cell get around the problem on the lagging strand? Once helicase has opened enough room to make it worthwhile, primase will put down a primer on the lagging strand (Figure 11.18, middle panel and homemade figure replication panel E) but please note that the 3 end of the primer is facing back toward the origin, and facing away from the region being opened up by helicase). DNA polymerase III will work, creating DNA on the lagging strand at the newly created primers 3 end of the RNA primer. (study guide compare figure E & F). But on the lagging strand synthesis will have to stop when polymerase runs into a previous primer Figure 11.18, bottom panel. Because on the lagging strand, the DNA polymerase III is always working in the backward direction (away from the direction the fork is opening), it will have to work in interrupted spurts. We describe this by saying DNA replication on the lagging strand is discontinuous. The short segments of DNA created on the lagging strand are called Okazaki fragments (figure 11.18, page 247 bottom panel). Step 7. The replication of the lagging strand takes a little longer but it does get done too. (study guide replication figure, part F, G, H). and text Figure 11.18, and 11.19, page 247. Now that we have made a lot of DNA, are we finished? The answer is no. Keep in mind the RNA primers must be removed! We can not keep RNA in our DNA polymers; RNA is not as stable. When DNA polymerase runs into an RNA primer, it comes off of the DNA and a special DNA polymerase, called DNA polymerase I comes in. Figure 11.19, page 247 between panels 2 and 3. Figure 11.19, page 247 second panel DNA polymerase I recognizes areas where RNA primer and DNA synthesis meet. Below red = RNA, black = DNA; arrow points to where they meet 5 3 3 5 Next, DNA polymerase I chews up the primer (one ribonucleotide at a time from the 5 end; this is referred to as its 5-3 exonuclease function). 5 3 3 5 www.notesolution.com
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