biology 1.3.PDF

Chapter 1: ilNA $tructurs, $ynthesis, and Repair -essan 1.3 DNA Repair DNA replication must be highly accurate to avoid introduction of mistakes into newly copied DNA strands. The error rate for DNA polymerases is estimated to be once every -105 nucleotides. One factor that :llows for such high...

Chapter 1: ilNA $tructurs, $ynthesis, and Repair -essan 1.3 DNA Repair DNA replication must be highly accurate to avoid introduction of mistakes into newly copied DNA strands. The error rate for DNA polymerases is estimated to be once every -105 nucleotides. One factor that :llows for such high accuracy is the nucleotide selectivity of DNA polymerases. DNA polymerases also -ave a proofreading function, which further improves the accuracy of replication by a factor of 100-1 ,000. - addition to their ability to add nucleotides to a growing strand, DNA polymerases possess exonuctease ::tivity, which allows them to remove bases from the end of a DNA strand. When a mismatch is detected :,rectly after replication or when DNA damage occurs outside of the replication process, distinctive DNA 'epair pathways are activated. This lesson discusses various mechanisms of DNA proofreading and ':cair. 1 ,3.01 DNA Proofreading and Mismatch Repalr I uring DNA replication, DNA polymerases may incorporate an incorrect nucleotide into the newly s:"nthesized strand, creating a Lrn$t,: llr,rit ii:irni;rl,:'ii between the daughter and parent strands (eg, G-T and r-C mispairing instead of proper G-C and A-T pairing). DNA polymerases are equipped with 3') 5' exonuclease activity, which acts as a proofreading function, allowing incorrectly paired nucleotides to be 'eplaced during DNA replication (Figure 1.16). DNA polymerase adds En inconect base......but detects the mistake and corrects it. Extension -W ry {t-*-'. Exonuclease Exonuclease domain activity Figure 1.16 DNA polymerases can correct nucleotide mismatches during replication. ren a base pair mismatch is detected immediately after replication, the DNA mismatch repair (MMR) ' stem is activated, as shown in Figure 1.17. ln MMR, an endonuclease enzyme removes (excises) the s"' -ismatched base and several nucleotides on either side of it from the daughter strand. ::ckaryotes can distinguish the template from the daughter strand by the state of strand rreti-rylatii:n. -re template strand has methyl groups attached to some of its bases, whereas the newly synthesized 17  Chapter 1: DNA Structure, $ynthesis, and Repair the daughter strand does not. In eukaryotes, the process of determining which strand contains breaks found only in newly synthesized DNA. mismatched base involves recognition of singte-stranded mismatched MMR enzymes must have endonuclease, rather than exonuclease, activity because can cleave nucleotides must be excised from within an existing DNA strand, and only endonucleases pfro*1:li*Lxiurlr,rr irilnil$ in the middle of a nucleic acid strand. After mismatched nucleotides are excised, brun potymerase incorporates appropriate nucleotides and DNA ligase catalyzes the formation of new phosphodiester bonds to rejoin the strands. New DNA Mismatch detected Endonuclease cleaves mispaired nucleotide and adjacent nucleotides Corrected nucleotides DNA polymerase adds correct nucleotides Sealed nick I DNA ligase seals nick Figure 1.17 Mismatch repair system' When both the inherent accuracy of DNA polymerase, its proofreading ability, and the MMR system are taken into account, the error rate of DNA replication is approximately one in every 1010-1011 nucleotides. ,18  Chapter 1: DNA Structure, Syilthesis, and Repair 1.3.02 DNA Damage and Repair Enors in replication are a common cause of DNA mismatches, but at other times during the cell cycle nucleic acids may also undergo spontaneous changes or damage due to exposure to harmful conditions (eg, UV light, high energy radiation, X-rays, chemicals). When DNA damage is detected, specific DNA rcpair pathways are activated, depending on the type of damage. The base excision repair system (see Figure 1.18) corrects damage that does not cause significant distortion of the double helix, such as oxidation, deamination, and alkylation of bases. A DNA glycosylase enzyme recognizes the site of DNA damage and excises the damaged base. An endonuclease enzyme then cleaves the phosphodiester bond, and DNA polymerase fills the gaps while DNA ligase rejoins the DNA strands. Dfimngadbassh&b# Enqymswifr Damqgsd ba$B ls Nxckad W Sugar4lmryhatr bed6one h obaved DI{A polymenra flls gapo and bllA llgne* aels nlcl$ Figure 1.18 Base excision repair system. lf damage to DNA is more extensive or bulky, the nucleotide excision repair (NER) pathway is activated, shown in Figure 1.19. For example, UV radiation can cause thymine dimers, or linkages of two thymine bases, which leads to a distortion in the double helix. When this type of damage is detected,  Chapter 1: DNA Strueture, Synthesis, and Repair DNA polymerase and DNA NER endonuclease enzymes are mobilized to remove the damaged region. ligase subsequently fill in the gaps and rejoin the repaired DNA strands. UV oxposure create$ thymine dimers Endonuclease clcavas DNAstrand and damaged DNA i$ discarded OllA polymonso adds new nucleo$das Sealed nlck Dt{A llga$€ soale nlck Figure 1.19 Nucleotide excision repair system' parts of the chromosome. lf these ln some situations, double-stranded breaks in DNA can separate breaks are not repaired, large portions of the chromosome can be lost. There are two major mechanism for repairing double-stranded breaks: homologous recombination and nonhomologous end joining' Thet two mechanisms are summarized in Figure 1.20' S and Gz phases of the cell lf an organism is diploid or a sister chromatid is nearby (such as during the chromosome, a cycle), the corresponding chromosome is used as a template to repair the broken is not present (eg, process known as nomiogous recombination. However, if a suitable homologue i strands can be rejoined by haploid organisms or when-no duplicated chromosome is present), broken involved in nonhomologous enzymes iia nonhomologous end joining. Because there is no template at the site where the DNA is rejoined' end joining, mutations arJoften incorporated 20  Chapter 1: DNA Structure, Synthesis, and Repair Homologous recombination Nonhomologour end folning Double'standed Double.stnanded breakin DNA break in DNA giffi,@3: i:@@3; W ry '$re3' l-*ornnlcgaus Broken ends rejolned by engyrnes Notemplate fmmS' shf0rnCIfisrns without a template; a ru w3' pmvidesGmplate 5'w wgn mutations may 1r m w5', lor repair $'mw wnru5f be inboduM W f, 5' 3' _*' 3 5' Remlred Repairsd drrunpsome i chrornosome 1.20 Homologous recombination and nonhomologous end joining. fr b.r  Chapter 1: DNA $tructure, Synthetiis, and Repair Lesson 1.4 Eukaryotic Chromosome Organization lntroduction In a typical eukaryotic cell, each linear chromosome consists of a DNA molecule that contains an average of 1.5 x 108 nucleotide pairs. A completely unwound chromosome would be about 4 cm long, which poses a problem because many chromosomes must fit into the nucleus of a cell. To address this problem, eukaryotic cells organize and condense DNA, packaging it densely with proteins to create chromatin, Chromatin packaging changes dramatically during cell division, and modification of chromatin structure is intricately linked with gene regulation. This lesson explores the organization of eukaryotic chromosomes and different states of chromosome organization. 1.4.01 Histones and Nucleosomes : During the initial steps of chromatin formation, the DNA double helix wraps around a complex of eight histone proteins (an octamer) two times to form a structural subunit called a nucleosome (Figure 1.21). There are five major histone types (ie, Hl ,H2A, H2B, H3, and H4). A DNA-wrapped octamer core is composed of eight subunits (two each of H2A, H2B, H3, and H4), and a linker histone (H1) is present outside of the core. Histone tail Linker histone Figure 1.21 Components of a nucleosome. Because DNA is negatively charged, histone proteins must have a net positive charge to facilitate DNA binding. The rich presence of arginine and lysine (positively charged, basic amino acidq) in histones confers this necessary positive charge. 23  Chapter 1: DNA Structure, $ynthesis, and Repair In unwound chromatin, the nucleosomes resemble beads on a string, and the DNA between each "bead" is called linker DNA (see Figure 1.22). During replication and transcription, when access to the DNA helix is required, the histones leave the DNA for a short period of time. Nucleosome core Acetyl.group (HzA, H2B, H3,l-14)2 Chromatin fiber Linker DNA Histone Hl (outside core) Chrcmosome Figure 1.22 Eukaryotic DNA organization. 1,4.02 Euchromatin and Heterochromatin When DNA is not actively being replicated or transcribed, chromatin may be configured either loosely (euchromatin) or densely (heterochromatin), as shown in Figure 1.23. Whether chromatin is configured as euchromatin or heterochromatin impacts its accessibility and the expression of genes located in that region of the chromosome. During cell division, chromatin becomes even more condensed in preparation for mitosis (see Concept5.4.02).  Chapter 1: DNA $tructure, $ynthebis, and Repair Figure 1.23 Euchromatin and heterochromatin. Heterochromatin consists of DNA tightly coiled around histone proteins, bound by ionic interactions between negatively charged phosphates on the DNA backbone and positively charged lysine residues on ttp histones. Because DNA in heterochromatin is densely packed, it is not readily accessible to hanscription machinery. Mafiy key noncoding regions of the chromosome, such as telomeres and centromeres, are composed of heteroch romatin. Euchromatin forms when hisiones are modified, often due to acetylation of lysine residues. Acetylation neutralizes the positive charge op the lysine residue, which reduces interactions between histones and DNA. These reduced interactionsyield a more open form of chromatin that allows better accessibility for banscription machinery. 25