DNA Replication PDF

During DNA replication, base pairing enables existing DNA strands to serve as templates for new complimentary strands  When a cell copies a DNA molecule, each strand serves as a template for ordering nucleotides into a new complimentary strand.   Nucleotides line up along the template strand a...

During DNA replication, base pairing enables existing DNA strands to serve as templates for new complimentary strands  When a cell copies a DNA molecule, each strand serves as a template for ordering nucleotides into a new complimentary strand.   Nucleotides line up along the template strand according to the base-pairing rules. The nucleotides are linked to form new strands (complementary). DNA Replication: 1. During DNA replication, base pairing enables existing DNA strands to serve as templates for new complimentary strands. 2. Several enzymes and other proteins carry out DNA replication: Helicase, Primase, Polymerase, Ligase. The ends of DNA molecules are replicated by a special mechanism. The Replication Mechanism  It takes E. coli less than an hour to copy each of the 5 million base pairs in its single chromosome and divide to form two identical daughter cells.  A human cell can copy its 6 billion base pairs and divide into daughter cells in only a few hours.  This process is remarkably accurate, with only one error per billion nucleotides.  A helicase; untwists and separates the template DNA strands at the replication fork.  Single-strand binding proteins; keep the unpaired template strands apart during replication. • The replication of a DNA molecule begins at special site called origin of replication which is a single specific sequence of nucleotides that is recognized by the replication enzymes. • Replication enzymes separate the strands, forming a replication “bubble”. – Replication proceeds in both directions until the entire molecule is copied.  In eukaryotes, there may be hundreds or thousands of bubbles (each has origin sites for replication) per chromosome.   At the origin sites, the DNA strands separate forming a replication “bubble” with replication forks at each end. The replication bubbles elongate as the DNA is replicated and eventually fuse.  Primer: (a short segment of RNA, 10 nucleotides long) is required to start a new chain.  Primase: (an RNA polymerase) links ribonucleotides that are complementary to the DNA template into the primer. • DNA polymerases: catalyze the elongation of new DNA at a replication fork. After formation of the primer, DNA polymerases can add deoxyribonucleotides to the 3’ end of the ribonucleotide chain. • Another DNA polymerase later replaces the primer ribonucleotides with deoxyribonucleotides complimentary to the template.  DNA polymerases can only add nucleotides to the free 3’ end of a growing DNA strand.  A new DNA strand can only elongate in the 5’->3’ direction.  At the replication fork, one parental strand (3’-> 5’ into the fork), the leading strand, can be used by polymerases as a template for a continuous complimentary strand. To elongate the other new strand of DNA in the obligatory 5’->3’ direction, DNA pol III must work along the other template strand in the direction away from the replication Fork . The DNA strand elongating in this direction is called the lagging strand. the lagging strand is synthesized discontinuously, as a series of segments (called Okazaki fragments. laying  Okazaki fragments (each about 100-200 nucleotides) are joined by DNA ligase to form the sugar-phosphate backbone of a single DNA strand. Step 1  Helicases: Enzymes that separate the DNA strands  Helicase move along the strands and breaks the hydrogen bonds between the complimentary nitrogen bases  Replication Fork: the Y shaped region that results from the separation of the strands Step 2 • DNA Polymerase: enzymes that add complimentary nucleotides. • Nucleotides are found floating freely inside the nucleus • Covalent bonds form between the phosphate group of one nucleotide and the deoxyribose of another • Hydrogen bonds form between the complimentary nitrogen bases Step 3 • DNA polymerases finish replicating the DNA and fall off. • The result is two identical DNA molecules that are ready to move to new cells in cell division. • Semi-Conservative Replication: this type of replication where one strand is from the original molecule and the other strand is new  Each strand is making its own new strand.  DNA synthesis is occurring in two different directions  One strand is being made towards the replication fork and the other is being made away from the fork. The strand being made away from the fork has gaps.  Gaps are later joined by another enzyme, DNA ligase • • The strands in the double helix are antiparallel. The sugar-phosphate backbones run in opposite directions. – Each DNA strand has a 3’ end with a free OH group attached to deoxyribose and a 5’ end with a free phosphate group attached to deoxyribose. – The 5’ -> 3’ direction of one strand runs counter to ‫ ُمعاكس لـ‬the 3’ -> 5’ direction of the other strand. SUMMARY OF DNA REPLICATION MECHANISM The two DNA-strands separate forming replication bubbles. Each strand functions as a template for synthesizing new complementary & lagging strands via primers, polymerase and ligase. 3 5 T A C T G A C A T G A C T G 3 5 Complementary (leading) strand T A C T G Primer Polymeras Ligase e A C 5 3 Lagging strand (complementary) Okazaki fragments Templates 1 2 3 4 Fig. 16.15, Page 298