1. Mutation The term mutation refers to both (1) the change in the genetic material and (2) the process by which the change occurs. An organism that exhibits a novel phenotype resulting from a mutation is called a mutant. Used in its broad historical sense. mutation refers to any sudden, heritable change in the genotype Reverse Mutation: When a second mutation restores the original phenotype lost because of an earlier mutation, the process is called reversion or reverse mutation. Reversion may occur in two different ways: (1) by back mutation, a second mutation at the same site in the gene as the original mutation, restoring the wild type nucleotide sequence, or (2) by the occurrence of a suppressor mutation, a second mutation at a different location in the

3. MUTATION: PHENOTYPIC EFFECTS The effects of mutations on phenotype range from alterations so minor that they can be detected only by special genetic or biochemical techniques, to gross modifications of morphology to lethals. Any mutation occurring within a given gene will thus produce a new allele of that gene. Genes containing mutations with no effect on phenotype or small effects that can be recognized only by special techniques are called isoalleles. Other mutations produce null alleles that result in no gene product or totally nonfunctional gene products. If mutations of the latter type occur in genes that are required for the growth of the organism, individuals that are homozygous for the mutation will not survive. Such mutations are called recessive lethals. Mutations can be either recessive or dominant. In monoploid organisms such as viruses and bacteria, both recessive and dominant mutations can be recognized by their effect on the phenotype of the organism in which they occur.

5. Mutations with Phenotypic Effects: Usually Deleterious and Recessive Most of the thousands of mutations that have been identified and studied by geneticists are deleterious and recessive. Effects of Mutations in Human Globin Genes The major form of hemoglobin in adults (hemoglobin A) contains two identical alpha (a) chains and two identical beta (13) chains. Each α polypeptide consists of a specific sequence of 141 amino acids, whereas each β chain is 146 amino acids long. Over 100 hemoglobin variants with amino acid chang the (3 chain are known. Most of them differ from the normal chain of hemoglobin A by a single amino acid substitution.

8. Mutation in Humans: Blocks in Metabolic Pathways Each step of metabolic pathway is catalyzed by an enzyme encoded by one or more genes. When mutations occur in such genes, they often cause metabolic blocks that lead to abnormal phenotypes. For example the metabolism of the aromatic amino acids phenylalanine and tyrosine provides an especially good example. Phenylalanine and tyrosine are essential amino acids required for protein synthesis. Ist-known inherited defect in phenylalanine-tyrosine metabolism is phenylketonuria, which is caused by the absence of the enzyme hydroxylase, the enzyme that converts phenylalanine to tyrosine. Newborns with phenylketonuria, an autosomal recessive disease, develop severe mental retardation if not placed on a diet low in phenylalanine

9. . The first inherited disorder in the phenylaanine-tyrosine metabolic pathway to be studied in humans was alkaptonuria, which is caused by autosomäl recessive mutations that inactivate the enzyme homogentisic acid oxidase. Tyrosinosis and tyrosinemia result from the lack of the enzymes tyrosine transaminase and p- hydroxyphenylpyruvic acid oxidase, respectively.. Individuals with tyrosinosis show pronounced increases in tyrosine levels in their blood and urine and have various congenital abnormalities. Individuals with tyrosinemia have elevated level of both tyrosine and p- hydroxyphenylpyruvic acid in their blood and urine. Most newborns with tyrosinemia die within six months after birth because of liver failure.

10. Albinism, the absence of pigmentation in the skin, hair, and results from a mutational block in the conversion of tyrosine the dark pigment melanin. One type of albinism is caused by the absence of tyrosinase, the enzyme that catalyzes the first step in the synthesis of melanin from tyrosine. Other types of albinism result from blocks in subsequent steps in the conversion of tyrosine to melanin. Albinism is inherited as an autosomal recessive trait; heterozygotes usually have normal levels of pigmentation. Therefore, two albinos who have mutations in different genes will produce normally pigmented children.

12. THE MOLECULAR BASIS OF MUTATION Mutations alter the nucleotide sequences of genes in several ways, for example the substitution of one base pair for another or the deletion or addition of one or a few base pairs. Watson and Crick pointed out that the structures of the bases in DNA are not static. Hydrogen atoms can move from one position in a purine or pyrimidine to another position—for example, from an amino group to a ring nitrogen. Such chemical fluctuations are called tautomeric shifts The more stable keto forms of thymine and guanine and the amino forms of adenine and cytosine may infrequently undergo tautomeric shifts to less stable enol and imino forms, respectively. The bases would be expected to exist in their less stable tautomeric forms for only short periods of time. However, if a base existed in the rare form at the moment that it was being replicated or being incorporated into a nascent DNA chain, a mutation would result. When the bases are present in their rare imino or enol states, they can form adenine-cytosine and guanine- thymine base pairs. The net effect such an event, and the subsequent replication required to separate the mismatched base pair, is an A:T to G:C or a G:C to 1: T base-pair substitution.

14. Transition and Transversion Mutations resulting from tautomeric shifts in the bases of DNA involve the replacement of a purine in one strand of DNA with the other purine and the replacement of a pyrimine the complementary strand with the other pyrimidine. Such base-pair substitutions are called transitions. Base- pair substitutions involving the replacement of a purine with a pvrimidine and vice versa are called transversions. Another type of point mutation involves the addition or deletion of one or a few base pairs. A surprisingly large proportion of the spontaneous mutations that have been studied are single base-pair additions and deletions rather than base-pair substitutions. These frameshift mutations almost always result in the synthesis of nonfunctional protein gene products.

17. Molecular mechanism of mutation Although much remains to be learned about the causes, molecular mechanisms, and frequency of spontaneously occurring mutations, three major factors are (1) the accuracy of the DNA replication machinery; (2) the efficiency of the mechanisms that have evolved for the repair of damaged DNA, and (3) the degree of exposure to mutagenic agents present in the environment. Perturbations of the DNA replication apparatus or DNA repair systems, both of which are under genetic control, have been shown to cause large increases in mutation rates.

18. Induced mutation Geneticists could now induce mutations in genes of interest and then study the effects of the missing gene products. We discuss Muller’s ingenious demonstration of X-ray-induced mutations on the X chromosome of Drosophila in A Milestone in Genetics: Muller DemonstratesThat X Rays Are Mutagenic. Subsequent work showed that X rays are mutagenic to all organisms and that many other agents— physical, chemical, and transposable genetic elements—are similarly mutagenic. X rays have many effects on living tissues. Mutagenic effects of mustard gas. A large class of chemical mutagens that transfer alkyl (CH, CH 3 CH, and so forth) groups to the bases in DNA; thus, they are called alkylating agents. Like X rays, mustard gas has many effects on

19. Mutations Induced by Radiation Ionizing radiations are of high energy and are useful forridical diagnosis because they penetrate living tissues for subitial distances. In the process, these high-energy rays collide with atoms and cause the release of electrons, creating positively charged free radicals or ions. The ions, in turn, collide with other molecules and cause the release of additional electrons. The result is that a cone of ions is formed along the track leach high-energy ray as it passes through living tissues. Ultraviolet rays, having lower energy than ionizing radiations, penetrate only the surface layer of cells in higher plants and animals and do not cause ionizations. Because of UV thymine-thymine dimer can be formed. For example, X-irradiation of Drosophila sperm causes an approximately 3 percent increase in mutation rate for each 1000 r increase in irradiation dosage.

20. Mutations Induced by Transposable Genetic Elements Living organisms contain remarkable DNA elements that can move from one site in the genome to another site. Mechanism of transposon- induced mutation. The insertion of a transposable genetic element (red) into a wild-type gene (left) will usually render the gene nonfunctional (right). A truncated gene product usually results from transcription- or translation- termination signals, or both, located within the transposon.

21. Expanding Trinucleotide Repeats and Inherited Human Diseases Repeated sequences of one to six nucleotide pairs are known as simple tandem repeats. Such repeats are dispersed throughout the human genome. Repeats of three nucleotide pairs, trinucleotide repeats, can increase in copy number and cause inherited diseases in humans. Several trinucleotides have been shown to undergo such increases in copy number. Expanded CGG trinucleotide repeats at the FRAXA site on the X chromosome are responsible for fragile X syndrome, the most common form of inherited mental retardation in humans ). Normal X chromosomes contain from 6 to about 50 copies of the CGG repeat at the FRAXA site. Mutant X chromosomes contain up to 1000 copies of the tandem CGG repeat at this site.

22. CAG and CTG trinucleotide repeats are involved in several inherited neurological diseases, including Huntington disease,, myotonic dystrophy, Kennedy disease, dentatorubral pallidoluysian atrophy, Machado-Joseph disease, and spinocerebellar ataxia. In all of these neurological disorders, the severity of the disease is correlated with trinucleotide copy number—the higher the copy number, the more severe the disease symptoms. In addition, the expanded trinucleotides associated with these diseases are unstable in somatic cells and between generations.

23. DNA REPAIR MECHANISMS Living organisms contain many enzymes that scan their DNA for damage and initiate repair processes when damage is detected.’ The multiplicity of repair mechanisms that have evolved in organisms ranging from bacteria to humans emphatically documents the importance of keeping mutation a tolerable level. For example, E. coli cells possess five well characterized mechanisms for the repair of defects in DNA: (1) light—dependent repair or photoreactivation, (2) excision repair, mismatch repair, (4) postreplication repair, and (5) the error repair system (SOS response). Moreover, there are at least two different types of excision repair, and the excision repair pathways be initiated by several different enzymes, each acting on specific kind of damage in DNA. Mammals seem to possess all the repair mechanisms found in E. coli except photoreactivation.

24. Light-Dependent Repair Light-dependent repair or photoreactivation of DNA in bacteria is carried Out by a light-activated enzyme called )DNA photolyase. When DNA is exposed to ultraviolet light, thymine dimers are produced by covalent cross linkages between adjacent thymine residues. DNA photolyase recognizes and binds to thymine dimers in DNA, and uses light energy to cleave the covalent cross- links. Photolyase will bind to thymine dimers in DNA in the dark, but it cannot catalyze cleavage of the bonds joining the thymine moieties without energy derived from visible light, specifically light within the blue region of the spectrum. Photolyase also splits cytosine dimers and cytosine-thymine dimers. Thus, when ultraviolet light is used to induce mutations in bacteria, the irradiated cells are grown in the dark for a few generations to maximize the mutation frequency.

26. Excision Repair Excision repair of damaged DNA involves at least three steps. In step 1, a DNA repair endonuclease or endonuclease- containing enzyme complex recognizes, binds to, and excises the damaged base or bases in DNA. In step 2, a DNA polymerase fills in the gap by using the undamaged complementary strand of DNA as template. In step 3, the enzyme DNA ligase seals the break left by DNA polymerase to complete the repair process. There are two major types of excision repair: base excision repair systems remove abnormal or chemically modified bases from DNA, whereas nucleotide excision repair pathways remove larger defects like thymine dimers. Both excision pathways are operative in the dark, and both occur by very similar mechanisms in E. coli and humans. Base excision repair can be initiated by any of a group of enzymes called DNA glycosylases that recognize

27. abnormal bases in DNA. Each glycosylase recognizes a specilk type of altered base, such as deaminated bases, oxidized bases.. and so on (step 2). The glycosylases cleave the glycosidic bo4 between the abnormal base and 2-deoxyribose, creatnf apurinic or apyrimidinic sites (AP sites) with missing bases (step 3). AP sites are recognized by enzymes called AP endonucleases, which act together with phosphodiesterases to excise the sugar- phosphate groups at these sites (step 4). DNA pokmerase then replaces the missing nucleotide according to specifications of the complementary strand (step 5), and DNA ligase seals the nick (step 6).

28. Nucleotide excision repair removes larger lesions like thymine dimers and bases with bulky side-groups from DNA. In nucleotide excision repair, a unique excision nuclease activity produces as other side of the damaged nucleotide(s) and excises an nucleotide containing the damaged base(s). In E. coli, exonuclease activity requires the production of three genes, uvrA, uvrB, and uvrC (designated uvr for ir). A trimeric protein containing two UvrA polypepone UvrB polypeptide recognizes the defect in DNA, binds to it, and uses energy from ATP to bend the DNA at the damaged site. The UvrA dimer is then released, and the UvrC protein binds to the UvrB/DNA complex. The UvrB protein cleaves the fifth phosphodiester bond from the damaged nucleotide(s) on the 3’ side, and the UvrC protein hydrolyzes the eighth phosphodiester linkage from the damage on the 5’ side. The uvrD gene product, DNA helicase II, releases the excised dodecamer. In the last two steps of the pathway, DNA polymerase I fills in the gap, and DNA ligase seals the remaining nick in the DNA molecule. Nucleotide excision repair in humans occurs through a pathway similar to the one in E. coil, but it involves about four times as many proteins. In humans, the excinuclease activity contains 15 polypeptides. Protein XPA (for xeroderma pigmentosum protein A) recognizes and binds to the damaged nucleotide(s) in DNA. It then recruits the other proteins required for excinuclease activity. In humans, the excised oligomer is 24 to 32 nucleotides long rather than the 12- mer removed in E. coil. The gap is filled in by either DNA polymerase or € in humans, and DNA ligase completes the job.

30. Mutation and Diseases

31. Some common genetic disorders DisorderMutat ion Chromo some Symptoms Cri du chat D5 Feeding problems because of difficulty swallowing and sucking., low birth weight and poor growth.low birth weight · severe cognitive, speech, and motor delays. ·behavioral problems such as hyperactivity aggression, tantrums, and repetitive movements. ·unusual facial features which may change over time. excessive drooling, constipation.drooling Cystic Fibrosis P7q Poor growth and poor weight gain, accumulation of thick, sticky mucus, frequent chest infections and coughing or shortness of breath. Males can be infertile due to congenital absence of the vas deferens. Symptoms often appear in infancy and childhood, such as bowel obstruction due to meconium ileus in newborn babies. As the child grows, he or she will need to exercise to release mucus in the alveoli.infertilecongenital absence of the vas deferens bowel obstructionmeconium ileus Hemo philia Prolonged bleeding and re-bleeding are the diagnostic symptoms of haemophilia. Internal bleeding is common in people with severe haemophilia and some individuals with moderate haemophilia. The most characteristic type of internal bleed is a joint bleed where blood enters into the joint spaces. This is most common with severe haemophiliacs and can occur spontaneously (without evident trauma). If not treated promptly, joint bleeds can lead to permanent joint damage and disfigurement.Internal bleedingjoint spaces

32. Disord- er Mutat -ion Chromo -some Symptoms Neurofibr- omatosis 17q/22q/ ? People with Neurofibromatosis can be affected in many different ways. There is a high incidence of learning disabilities in people with NF. It is believed that at least 50% of people with NF have learning disabilities of some type. Increased chances of development of petit mal epilepsy (a Partial absence seizurepetit mal epilepsy disorder). The tumors that occur can grow anywhere a nerve is present. This means that: They can grow in places that are very visible. The tumors can also grow in places that can cause other medical issues that may require them to be removed for the patient's safety. Affected individuals may need multiple surgeries, depending on where the tumors are located. Tay-Sachs disease P15 It causes a relentless deterioration of mental and physical abilities that commences around six months of age and usually results in death by the age of four Duchenne muscular dystrophy DXpProgressive muscular wasting, Poor balance, Frequent falls Walking difficulty, Calf deformation,Limited range of Movement, Respiratory difficulty, Drooping eyelids Gonadal, Loss of bladder control, Scoliosis (curvature of the spine and the back), Inability to walkScoliosis Bladder cancer Bladder cancer characteristically causes blood in the urine; this may be visible to the naked eye (gross hematuria) or detectable only by microscope (microscopic hematuria). Other possible symptoms include pain during urination, frequent urination (polyuria) or feeling the need to urinate without results.hematuria pain during urinationpolyuria