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Biochemistry-II - DNA Structure, Replication, and Repair

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    Study Guide Biochemistry - II – DNA Structure, Replication, and Repair 1. DNA Repair Why DNA Repair Is Necessary DNA carries genetic information, but it is still a chemical molecule , and like all chemicals, it can be damaged by reactions. Any change that alters DNA sequence information is called a mutation . Substances or agents that cause mutations are known as mutagens . Without repair systems, mutations would accumulate so rapidly that life could not be maintained across generations. Types of DNA Mutations Several kinds of mutations occur, depending on how the DNA sequence is changed. Figure 1 1. Transition Mutations A transition is a substitution within the same base class: • A – T ↔ G – C

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    Study Guide Example: • Hydroxylamine removes the amino group from deoxycytosine • Deoxycytosine becomes deoxyuracil • During replication, uracil pairs with adenine • This converts G – C → A – T Hydroxylamine is a monodirectional mutagen because it causes only one type of base change. 2. Transversion Mutations A transversion occurs when: • A purine is replaced by a pyrimidine , or vice versa Examples: • A – T → T – A • A – T → C – G Ultraviolet (UV) light can cause transversions, though it can cause other mutations as well. 3. Frameshift Mutations Frameshift mutations occur when: • A base pair is inserted or deleted This shifts the reading frame of the gene, often producing a nonfunctional protein . Frameshifts are commonly caused by planar intercalating agents , which insert between DNA bases. Large insertions and deletions also occur naturally. Ultraviolet Light as a Mutagen UV radiation is a major environmental mutagen.

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    Study Guide • The ozone layer absorbs much UV radiation • Depletion of ozone (by CFCs) increases mutation risk All DNA bases absorb UV light, becoming chemically reactive. Pyrimidine Dimers UV light causes adjacent pyrimidines (especially thymine) to form pyrimidine dimers . • These dimers distort DNA structure • They block DNA replication and transcription

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    Study Guide Figure 2 Spontaneous Mutations Not all mutations are caused by external agents.

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    Study Guide Spontaneous mutations arise from: • Errors during DNA replication • Normal chemical reactions • Heat (since higher temperature increases reaction rates) Thus, even normal cellular conditions can cause mutations. DNA Repair Systems Cells contain many DNA repair mechanisms to counteract mutations. • Typical mutation rates are less than 1 per billion bases per generation • Humans have ~3 billion base pairs, so even identical twins differ by a few mutations • Viruses usually lack repair systems and rely on rapid replication instead Direct Reversal of DNA Damage Some damage can be reversed directly. Example: • Alkylating agents add methyl groups to guanine • Specialized repair proteins remove the methyl group • The repair protein is then degraded This restores the original DNA structure. Photoreactivation Repair UV - induced thymine dimers can be repaired directly by photoreactivation . • The enzyme DNA photolyase binds to thymine dimers • Visible light activates the enzyme • The dimer is split back into normal bases This is a direct and efficient repair method.

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    Study Guide Excision Repair: A General Repair System Many DNA lesions are repaired by excision repair . Key steps: 1. Damage is recognized (often by altered DNA shape) 2. A nuclease cuts the DNA near the lesion 3. The damaged segment is removed 4. DNA polymerase I fills in the missing bases 5. DNA ligase seals the strand The intact complementary strand provides the correct sequence. Repair of Deoxyuridine in DNA Special systems remove deoxyuridine (dU) from DNA.

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    Study Guide Figure 3 Sources of Deoxyuridine in DNA dU can appear in DNA due to: • Incorporation of dUTP during replication • Deamination of deoxycytidine If dU remains in DNA:

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    Study Guide • It pairs with adenine • This converts G – C → A – T after replication How Cells Prevent dU - Induced Mutations Cells use two main strategies: 1. dUTPase o Converts dUTP → dUMP o Prevents dUTP from being used by DNA polymerase 2. Base - excision repair o Removes uracil from DNA before replication Why DNA Uses Thymine Instead of Uracil Both thymine (T) and uracil (U) pair with adenine using two hydrogen bonds . However, thymine has a methyl group , while uracil does not. This difference is crucial: • If cytosine deaminates → uracil → easily recognized and repaired • If 5 - methylcytosine deaminates → thymine → hard to detect As a result: • Repair systems cannot tell which strand is wrong • These sites become mutation hot spots DNA methylation therefore increases mutation risk at certain sequences. Key Takeaway • DNA damage leads to mutations • Mutagens include chemicals, UV light, and heat • Major mutation types: transition, transversion, frameshift

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    Study Guide • UV light causes pyrimidine dimers • Cells use direct repair, photorepair , and excision repair • Special systems remove uracil from DNA • DNA uses thymine to improve mutation detection • Methylated DNA regions are mutation hot spots 2. DNA Recombination and Repair When DNA Damage Is Severe Sometimes DNA damage is so extensive that normal repair systems are not enough. In these situations, cells activate emergency mechanisms to preserve genetic information and survive . Two important strategies are: • SOS response (error - prone repair) • Recombinational repair Both aim to keep the chromosome functional, even under extreme stress. 2.1 The SOS Response (Error - Prone Repair) The SOS response is a last - resort repair system. • It is activated when DNA damage is severe • The goal is to allow replication to continue • Accuracy is sacrificed for survival • Mutations are more likely, which is why it is called error - prone repair Although imperfect, this response can prevent complete loss of genetic information. Recombinational Repair: Using Backup Information Recombinational repair allows one DNA copy to repair another copy during replication.

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    Study Guide • Occurs at the replication fork • One daughter chromosome supplies correct information to the other • Ensures that overall genetic information remains intact This system relies heavily on homologous recombination . Role of the RecA Protein The key protein in bacterial recombination and repair is RecA . • “Rec” stands for recombination • Cells lacking RecA cannot recombine DNA • Such cells are extremely sensitive to UV radiation RecA is a strand - exchange protein that initiates recombination.

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