Chapter 4Gene Function

Copyright © 2010 Pearson Education Inc.

Genes encode proteins, including enzymes. Genes work in sets to accomplish biochemical

pathways. Genes often work in cooperation with other

genes. These discoveries are the foundation of modern

molecular genetics.

Alkaptonuria is a human trait characterized by urine blackening on exposure to air and arthritis in later life.

Archibald Garrod and William Bateson (1902) concluded alkaptonuria is genetically determined because:◦ a. Families with alkaptonuria often have

several affected members.◦ b. Alkaptonuria is much more common in first-

cousin marriages than marriages with unrelated partners.

Garrod showed that alkaptonuria results from homogentisic acid (HA) in the urine. Garrod reasoned that normal people metabolize HA, but those with alkaptonuria do not because they lack the necessary enzyme. He termed this an inborn error of metabolism .

The responsible mutation is recessive. The gene was later shown to be on chromosome 3.

Genes act by regulating definite chemical events.

George Beadle and Edward Tatum (1942) showed a direct relationship between genes and enzymes in the haploid fungus Neurospora crassa. This led to their one-gene–one-enzyme hypothesis, and a share of the 1958 Nobel Prize in Physiology or Medicine.

Neurospora propagates asexually by dispersal of bits of mycelium or by conidia.◦ b. It also propagates

sexually by means of two mating types, a and a.

◦ c. Wild-type Neurospora needs only simple minimal media with: i.Inorganic salts

(including a nitrogen source). ii.An organic carbon

source (such as glucose or sucrose).

iii.Biotin (a vitamin).

Beadle and Tatum isolated auxotrophic mutants by mutating with X-rays and then crossing with a wild-type strain. The cross ensured that effects were due to inheritance, rather than to direct damage from the radiation. In their experiment:◦ a. One progeny spore per

ascus was germinated in a complete medium so that growth would occur regardless of nutritional mutations. Then cells were transferred to minimal media, where auxotrophs won’t grow.

Each mutant was then tested on an array of minimal media, each with a different single supplement, to determine the type of nutritional mutation.

Reasoning: metabolism proceeds by a series of reactions, each catalyzed by an enzyme, and organized into pathways. Methionine biosynthesis as example◦ a. started with a set of methionine auxotrophs and found that

four genes are involved: met-2+, met-3+, met-5+, met-8+.◦ b. Check each mutant on a series of minimal media, each

supplemented with a different chemical believed to be involved in the pathway.

◦ c. They were able to deduce the pathway of methionine synthesis and to correlate mutations with enzymes used in the pathway.

Beadle and Tatum’s famous conclusion from this experiment is that one gene encodes one enzyme. Later work showed that some proteins consist of more than one polypeptide, and that not all proteins are enzymes. The principle is now usually stated as one-gene–one-polypeptide.

The enzymes involved in biochemical pathways within cells are under genetic control. The sum of all chemical intermediates and products in these biochemical pathways is the cell’s metabolome. Metabolomics is the study of the metabolome.

Single gene mutations are responsible for many human genetic diseases. Some mutations create a simple phenotype, while others are pleiotropic.

Phenylketonuria (PKU) is commonly caused by a mutation on chromosome 12 in the phenylalanine hydrolase gene.

Phenylalanine is an essential amino acid, excess is harmful and is normally converted to tyrosine. Excess phenylalanine affects the CNS, causing mental retardation, slow growth, and early death.

PKU’s effect is pleiotropic. Symptoms result from excess phenylalanine and from inability to make tyrosine; retardation,fair skin and blue eyes (even with brown-eye genes) and low adrenaline levels.

Diet: controlled intake of phenylalanine for protein synthesis but not enough that it accumulates.

The special diet must commence in the first two months after birth, continue at least throughout childhood,

PKU women have to resume the diet before pregnancy in to avoid phenylalanine levels that would affect the fetus.

NO Nutra-Sweet for all PKU individuals (Aspartam is aspartic-acid-phenylalanine)

Classic albinism is an autosomal recessive mutation in the gene for tyrosinase.

Tyrosinase converts tyrosine to DOPA in the melanin pathway.

Without melanin, individuals have white skin and hair, and red eyes, due to lack of pigmentation in the iris.

Two other forms of albinism are known, resulting from defects in other genes in the melanin pathway. A cross between parents with different forms of albinism can produce normal children.

Several genes can be mutated to cause this autosomal recessive disease

Mutated genes code parts of dynein motors of flagella and cilia, which slide the microtubules to produce motion.

Pleiotrophic effect:◦ a. Sinus and lung abnormalities. Due to loss of cilia

function, respiratory diseases are common.◦ b. Sterility. Sperm cannot swim and cilia cannot move the

oocyte within the reproductive tract.◦ c. Dextrocardia (heart shifted to the right), in some cases.

Cilia are involved in orientation of the embryonic heart.

The HexA enzyme cleaves a terminal N-acetylgalactosamine group from a brain ganglioside.

Tay–Sachs is one of a group of diseases called lysosomal-storage diseases.

Recessive mutations in genes encoding lysosomal enzymes.

Tay–Sachs disease (infantile amaurotic idiocy) recessive mutation in the gene hexA, which encodes the enzyme N-acetylhexosaminidase A.

◦ a. Enhanced reaction to sharp sounds.

◦ b. A cherry-colored spot surrounded by a white halo may be visible on the retina.

◦ c. Rapid neurological degeneration begins about age 1, as brain accumulate of unprocessed ganglioside.

◦ d. Blindness, hearing loss, and serious feeding problems lead to immobility by age 2.

◦ e. Death often occurs at 3 to 4 years of age, often from respiratory infection.

The disease is incurable. Carriers and affected individuals can be detected by genetic testing.

Infants will have nonfunctional HexA enzyme and the unprocessed ganglioside accumulates in brain cells causing various clinical symptoms:

Genes also make proteins that are not enzymes. Structural proteins, such as hemoglobin, are often abundant, making them easier to isolate and purify.

J. Herrick (1910) first described sickle-cell anemia. ◦ found that red blood cells (RBCs) change shape (form a

sickle) under low O2 tension

◦ a. Sickled RBCs are fragile, hence the anemia.◦ b. Less flexible than normal RBCs, forming blocks in

capillaries.◦ c. Pleiotropic, damage to extremities, heart, lungs,

brain, kidneys, GI tract, muscles, and joints. Others are: heart failure, pneumonia, paralysis, kidney failure, abdominal pain, and rheumatism.

◦ d. Heterozygous individuals have sickle-cell trait, a much milder form of the disease.

◦ a. Sickle-cell hemoglobin (Hb-S) has altered mobility compared with normal hemoglobin (Hb-A).

◦ b. Sickle-cell trait shows equal amounts of Hb-A and Hb-S, heterozygotes make both forms of hemoglobin.

◦ c. The sickle-cell mutation changes the form of its corresponding protein, and protein structure is controlled by genes.

Linus Pauling and his colleagues (1949) used electrophoresis and showed:

Hemoglobin has four polypeptide chains: two α polypeptide and two of the β polypeptide, each associated with a heme group.

V. M. Ingram (1956) found that the sixth amino acid of the β chain in sickle-cell hemoglobin is valine (no electrical charge) replacing the negatively charged glutamic acid of normal hemoglobin.

Outline of sickle-cell anemia and trait:◦ a. Wild-type β chain allele is A, which is codominant with S.◦ b. Hemoglobin of A/A individuals has normal subunits,

hemoglobin of genotype S/S has subunits that sickle at low O2 tension.

◦ c. Hemoglobin of A/S individuals is 1⁄2 normal and 1⁄2 sickling form. (The twochains of an individual hemoglobin molecule will be of the same type, rather than mixed.) Sickle-cell symptoms after sharp drop in the oxygen content of their environment.

One-gene–one-polypeptide hypothesis is a simplification. Alternative splicing in eukaryotes can result in more than one polypeptide from a single gene.

Screening of hemoglobin for altered electrophoretic mobility identified over 200 hemoglobin mutants.

Most effects are not as severe as those seen in sickle-cell anemia.

Cystic fibrosis (CF) is a recessive disease with peliotrophic effects:◦ affects the pancreas, lungs, and digestive system, and

sometimes the vas deferens in males. ◦ abnormally viscous secreted mucus resulting in lung

complications.◦ Life expectancy with current treatments is about 40 years

The gene is on the long arm of chromosome 7. Encodes a protein called cystic fibrosis

transmembrane conductance regulator (CFTR). Most common is 3bp deletion at position 508.

The structure of the protein has been deduced from its sequence. CFTR has homology with a large family of active transport membrane proteins.

Functional analysis shows that CFTR normally forms a chloride channel in the cell membrane. The mutation prevents chlorine ion transport and resulting in CF symptoms.

Mice genetically engineered to have the same defect in their CFTR gene are an animal model for the disease.

Genetic testing for a disease or carrier status, and chromosomal abnormalities.

The advice is based on genetic analysis, explaining diseases, probabilities, and options to affected individuals or parents.

Effective genetic counseling requires up-to-the-minute knowledge and is based on:◦ a. Pedigree analysis is an important tool of genetic

counseling, considering phenotypes found in both families over several generations. This is particularly useful for identifying suspected carriers.

◦ b. Fetal analysis includes assays for enzyme activity or protein level, or detection of changes in the DNA itself.

Prenatal DiagnosesPrenatal Diagnoses