1. CHAPTER 4 Gene Function Authored by Peter J. Russell

2. Gene Control of Enzyme Structure Genes encode proteins including enzymes, which catalyze reactions Genes work in sets to accomplish biochemical pathways Genes often work in cooperation with other genes

3. Garrod and Bateson’s Hypothesis of Inborn Errors of Metabolism – work in Alkaptonuria Alkaptonuria is symptomized by blackened, oxidized urine and late onset arthritis The etiology of the disease (genetic) was elucidated by examining familial inheritance Pathologically, patients with alkaptonuria lack the necessary enzyme to metabolize homogenistic acid (HA) due to a recessive mutation on chromosome 3 (found in studies performed later on) Garrod’s work provided the first evidence of a specific relationship between genes and enzymes.

4. Notice the relationship of pathways within the metabolome

5. Next, George Beadle and Edward Tatum exposed a bread mold (haploid fungus Neurospora crassa) to X-rays, creating mutants unable to survive on minimal medium due to an inability to synthesize certain molecules Using crosses, they identified three classes of arginine-deficient mutants, each lacking a different enzyme necessary for synthesizing arginine They developed a one-gene one-enzyme hypothesis, stating that each gene dictates the production of a specific enzyme One-Gene One-Enzyme Hypothesis Animation: Meselson-Stahl Experiment Animation: Meselson-Stahl Experiment

6. Life Cycle of Neurospora Asexual propogation occurs through the duplication of condidia or mycelium (haploid genome – mitotic) 1 Sexual reproduction occurs through the fusion of two mating types producing ascospores (diploid genome – meiotic) 2 Prototrophs (wild-type) grow on minimal medium (precursors only) Auxotrophs (mutants) need nutritional medium supplements (products) 11 2

7. Experimental Protocol One mating type of conidia were mutagenized with X-rays and crossed with wild- type conidia of the opposite mating type (Why?) Fruiting bodies were produced; microscopic ascospores were dissected and transferred to culture tubes containing complete or minimal medium No growth on minimal medium identified a nutritional mutant that was further investigated and specifically identified

8. Mutants affirm that methionine biosynthesis proceeds through a series of reactions catalyzed by enymes

9. But some proteins aren’t enzymes. So what then? Thus, the researchers revised the hypothesis to: one-gene one-protein But, But! Many proteins are composed of several polypeptides, each of which has its own gene Therefore, Beadle and Tatum’s hypothesis is now restated as the one-gene one- polypeptide hypothesis Note that even though it is not accurate (or sometimes correct) it is common to refer to gene products as proteins rather than polypeptides

10. Genetically based enzyme deficiencies

11. Presentation Projects Each of you will be required to present a 15 minute slide presentation (overheads in powerpoint) of a genetically based enzyme deficiency The format of the presentation will be like my presentation of Fibrodysplasia Ossificans Progressiva (a genetically based developmental defect) Your information resources are unlimited and some are included in my presentation for FOP For the NCBI database, go to http://omim.orghttp://omim.org Put in the OMIM entry number (on previous slide) for the genetic disorder you drew Pull up the information and construct your presentation from what is given

12. Gene Control of Protein Structure Genes also encode proteins that are not enzymes Structural proteins, such as hemoglobin, are often abundant, making them easier to isolate and purify (enzymes are generally produced in much smaller amounts and thus, are more difficult to purify to homogeneity) Animation: Gene control of Protein Structure and Function Animation: Gene control of Protein Structure and Function

13. Sickle Cell Anemia and Hemoglobin Hemoglobin is formed by four polypeptide chains and is responsible for O 2 and CO 2 distribution Two subunits of the protein contain the  polypeptide Two subunits of the protein contain the  polypeptide Each subunit associates with a heme group, which contains an iron center (reactive site)

14. Sickle Cell Hemoglobin Phenotypically, the mutant red blood cells change shape (sickle) under low O 2 tension Sickled cells are fragile, causing anemia. They are also less flexible, blocking up capillaries Effects are pleiotropic, including damage to the extremities, resulting in heart failure, pneumonia, paralysis, kidney failure, abdominal pain, and rheumatism Heterozygous individuals have the sickle-cell trait, a much milder form of the disease

15. Hemoglobin Form Electrophoresis of the protein showed the sickle cell form of Hb (Hb-S) has altered mobility compared with normal hemoglobin (Hb-A) Hemoglobin from individuals with the sickle-cell trait shows equal amounts of Hb-A and Hb-S, indicating that heterozygotes make both forms of Hb Thus, the sickle-cell mutation changes the form of its corresponding protein. Since protein structure is controlled by genes…

16. Hemoglobin Genetics The 6 th amino acid of the  chain in sickle-cell hemoglobin is valine (no electrical charge) rather than the negatively charged glutamic acid in the  chain of normal hemoglobin Over 200 types of hemoglobin mutants have been genetically characterized (most are point mutants) The mutant form is codominantly expressed

17. Cystic Fibrosis  The affected gene is on the long arm of chromosome 7 and encodes a protein called Cystic Fibrosis Transmembrane Conductance Regulator  Mutation results in an abnormal CFTR protein, preventing chlorine ion transport, resulting in mucus accumulation  Cystic fibrosis (CF) affects the pancreas, lungs, and digestive system, and sometimes the vas deferens in males  Treatment regimen includes antibiotics and percussion of the thoracic cavity

18. Genetic counseling Genetic counseling is advice based on genetic analysis, focusing either on the probability that an individual has a genetic defect or the probability that prospective parents will produce a child with a genetic defect Information for genetic counseling is obtained from two sources:  Pedigree Analysis (prior history of disease in a family?)  Carrier Detection (identify genotype of parents - Aa?)  Fetal Analysis (fetal biochemical or genetic assay)

19. Fetal Analysis Amniocentosis  Involves removal of a sample of amniotic fluid using a syringe needle inserted through the uterine wall (post 12 wk)  Fetal cells are cultured  Biochemical and genetic analyses are performed

20. Chorionic villus sampling  Involves removal of chorionic villus tissue either through the abdomen or vagina (wk 8-12)  No need for further culture of fetal cells  Biochemical and genetic analyses are performed

21. Amniocentesis is costly and cannot be performed until the second trimester, eliminating early abortion as an option in cases of severe genetic defect Chorionic villus sampling can be done earlier, but it carries a higher risk of fetal death and inaccurate diagnosis due to the presence of maternal cells Caveats