7. 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 Types of RNAs • mRNA: is the carrier of the genetic “message” from the DNA to the cytosol. • rRNA: is the major component of ribosomes. • tRNA: is the carrier of specific amino acids from the cytosol to ribosimes thus help to form polypeptides. RNA transcription and translation are the two main processing that link gene to protein • The information content of DNA is in the form of specific sequences of nucleotides along the DNA strands. • The DNA inherited by an organism leads to specific traits by dictating the synthesis of proteins. • Proteins are the links between genotype and phenotype. – For example, Mendel’s dwarf pea plants lack a functioning copy of the gene that specifies the synthesis of gibberellins (which stimulate the normal elongation of stems). • Genes provide the instructions for making specific proteins. • The bridge between DNA and protein synthesis is RNA. • The specific sequence of hundreds or thousands of nucleotides in each gene carries the information for the primary structure of a protein, the linear order of the 20 possible amino acids.  During RNA transcription, a DNA strand provides a template for the synthesis of a complementary RNA strand.  Transcription of a gene produces a messenger RNA (mRNA) molecule.  During RNA translation (at ribosomes), the information contained in the order of nucleotides in mRNA is used to determine the amino acid sequence of a polypeptide.  The basic mechanics of transcription and translation are similar in eukaryotes and prokaryotes.  Because bacteria lack nuclei, transcription and translation are coupled.  Ribosomes attach to the leading end of a mRNA molecule while transcription is still in progress. • In a eukaryotic cell, all transcription occurs in the nucleus and translation occurs mainly at ribosomes in the cytoplasm. • In addition, before the primary transcript can leave the nucleus it is modified in various ways during RNA processing before the finished mRNA go to the cytoplasm. • To summarize, genes program protein synthesis via genetic messenger RNA. • The molecular chain of command in a cell is : DNA Transcription mRNA Translation Protein In the genetic code, nucleotide triplets specify amino acids • • • • • • Triplets of nucleotide bases are the smallest units that can code for all the amino acid. In the triplet code three consecutive bases specify an amino acid. The genetic instructions for a polypeptide chain are written in DNA as a series of three-nucleotide words (triplets). During transcription, one DNA strand (the template strand) provides an RNA template. The complementary RNA molecule is synthesized according to base-pairing rules, except that uracil is the complementary base to adenine. During translation, blocks of three nucleotide bases (codons), are decoded ‫ فك الشفرة‬into a sequence of amino acids. • read in the 5’->3’ direction along the mRNA. • The codon UUU coded for the amino acid phenylalanine. • The codon AUG not only codes for the amino acid methionine but also indicates the start of translation. • A specific codon indicates a specific corresponding amino acid, but the amino acid may be the translation of several possible codons. The reading frame and subsequent codons are read in groups of three nucleotide bases (codon). Ay • stop start A During translation, the codons are • In summary, genetic information is encoded as a sequence of base triplets (codons) which is translated into a specific amino acid during protein synthesis. A  Transcription can be separated into three stages: 1- initiation 2- elongation, 3- termination.  Promotor contains the starting point for the transcription of a gene.  Promotor also includes a binding site for RNA polymerase.  Thus, RNA- polymerase can recognize and bind directly to the promotor region.  As RNA polymerase moves along the DNA, it untwists the double helix, 10 to 20 bases at time.  The enzyme adds nucleotides to the 3’ end of the growing strand.  Behind the point of RNA synthesis, the double helix reforms and the RNA molecule moves away.  Transcription proceeds until after the RNA polymerase transcribes a terminator sequence in the DNA. Eukaryotic cells modify RNA after transcription Enzymes in the eukaryotic nucleus modify pre-mRNA before the genetic messages are dispatched to the cytoplasm. 1)At the 5’ end of the pre-mRNA molecule, a modified form of guanine is added, the 5’ cap which function as: 1) 2) protect mRNA from hydrolytic ‫ ُمحلل‬enzymes. a translation start point for ribosomes. Eukaryotic cells modify RNA after transcription 2) At the 3’ end, an enzyme adds 50 to 250 adenine nucleotides, the poly(A) tail. The poly(A) tail facilitate the export of mRNA from the nucleus. 3) The removal of large portions of the RNA molecule that is initially synthesized (This cut-and-paste job, called RNA splicing)