Download Lecture 4 Gene Products

CHAPTER 4
Gene Function
• There is a specific relationship between
genes and enzymes, initially embodied in
the one gene-one polypeptide hypothesis.
• Since some enzymes consist of more than
one polypeptide chain and each gene
codifies for only one polypeptide chain, the
hypothesis became one gene-one
polypeptide.
• Many human diseases are caused by enzyme
deficiency (usually recessive traits).
• There is a large amount of evidence that
genes control the structure of all proteins.
A brief review of protein structure.
• The building blocks of proteins are amino
acids.
• An amino acid is a molecule that has an
carboxyl group and an amino group. R
represents the radical or rest of the
molecule.
Amino
group
Carboxyl (acid)
group
There are 20
different amino
acids. These can be
non-polar
(hydrophobic),
polar and
electrically charged.
Those that are
electrically charged
can be acidic (2
carboxyl groups)
or basic (2 amino
groups)
• Cells link amino
acids together by
dehydration
synthesis
• The bonds between
amino acid
monomers are
called peptide bonds
A protein’s primary structure is its amino acid
sequence.
Secondary structure is polypeptide coiling or
folding produced by hydrogen bonding.
Tertiary structure is the overall shape of a
polypeptide.
Quaternary structure is the relationship among
multiple polypeptides of a protein
Primary
structure
Amino
acid
Secondary
structure
Hydrogen
bond
Alpha
helix
Pleated
sheet
Tertiary
structure
Polypeptide
(single subunit
of transthyretin)
Quaternary
structure
Transthyretin, with four
identical polypeptide subunits
Garrod’s hypothesis of Inborn errors in
Metabolism.
ƒ In 1902 Archibald Garrod studied Alkaptonuria,
an inherited condition that causes urine to turn
black when exposed to air. Ochronosis, a
buildup of dark pigment in connective tissues
such as cartilage and skin, is also characteristic
of the disorder. People with alkaptonuria
typically develop arthritis in adulthood.
ƒ 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. HA is absent from normal 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 (Figure 4.1).
• Garrod’s work was the first evidence of a specific
relationship between genes and enzymes. He also studied
three other human diseases and provided further evidence.
Its most important contribution was the insight that a
mutation can block a human metabolic pathway by
damaging an enzyme, causing a detectable buildup of that
enzyme’s substrate.
*
* Phenyalanine
and Tyrosine
are amino
acids found in
the dietary
protein
*
Not found in
normal urine.
It turns black
in contact
with air.
How common is alkaptonuria?
• The condition is rare, affecting 1 in 250,000 to 1 million
people worldwide. Alkaptonuria is more common in certain
areas of Slovakia (where it has an incidence of about 1 in
19,000 people) and in the Dominican Republic.
What genes are related to alkaptonuria?
• Mutations in the HGD gene prevent the synthesis of the
enzyme that catalyzes the breakdown of homogentisic acid.
How do people inherit alkaptonuria?
• This condition is inherited in an autosomal recessive pattern.
Most often, the parents of an individual with an autosomal
recessive disorder are carriers (heterozygous) and do not show
signs and symptoms of the disorder.
The one-gene-one enzyme
hypothesis
• In 1942 Beadle and Tatum leaded the
beginning of biochemical genetics, a
branch of genetics that combines both
disciplines to explain the nature of
metabolic pathways.
• Beadle and Tatum used a haploid fungus,
Neurospora crassa (the orange mold that
grows on bread).
Neurospora as a Model organism.
• The mold Neurospora has a life cycle well suited to its use as a
model organism in genetics. First of all, the organism spends
most of its life cycle as a haploid organism. This means it is
possible to study the expression of genes without worrying
about dominance or recessive alleles.
• The fungus has alternate mating strains, here called type A and
type a. Mating can only take place between different mating
strains and the result is a diploid cell in a long sac.
• Because of the ascus the results of segregation during metaphase
1 are kept in order.
• Another advantage of neurospora is that in addition to
ascospores, the fungus also produces asexual spores
that allow scientists to clone any interesting Neurospora
genotypes.
• Next, the life cycle of Neurospora, is quite rapid
requiring about 2 weeks, allowing scientists to rapidly
conduct experiments.
• Wild-type Neurospora needs only simple minimal media
with:
– Inorganic salts (including a nitrogen source).
– An organic carbon source (such as glucose or
sucrose).
– Biotin (a vitamin).
• To grow on minimal media, wild-type Neurospora synthesizes
all organic molecules it needs for growth. However if a
mutation is introduced that impedes the production of one of
these molecules, it will not grow unless the molecule is added
to the minimal media . Organisms that grow in minimal
media are prototrophs.
• Auxotrophs or auxotrophic mutant are the type of mutant
that are unable to make a needed nutrient and will grow only if
the growth media is supplemented with the nutrient.
• A complete growth medium includes all 20 amino acids. Most
nutritional mutants can survive in this media.
• Wild type conidia (asexual spores) were irradiated with x rays.
After irradiation, they were germinated and cultured on maximal
media so that all Neurospora carrying possible mutations resulting
from the x rays would grow.
• These irradiated Neurospora were crossed with wild type
Neurospora to produce asci containing segregated products of
meiosis.
• The ascosposres were then isolated and grown on complete
media. Many hundreds of tubes are used for this step.
• Once the cultures were mature, asexual spores were isolated
from each tube and grown on minimal medium. Failure of a
specific spore to grow on minimum medium indicates the
presence of a mutant unable to synthesize a required compound
from the raw materials in the minimum medium.
Each mutant was then
tested on an array of
minimal media, each
with a different single
supplement, to
determine the type of
nutritional mutation
(Figure 4.2).
They started with a set of methionine auxotrophs, and found
that four genes are involved, met-2+, met-3+, met-5+, met-8+ .
• They checked each of these mutants on a series of
minimal media, each supplemented with a different
chemical believed to be involved in the pathway. They
expected growth if providing a chemical used after the
metabolic block, so the earlier the mutated gene
functions in the pathway, the more supplements will
support growth.
• 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.
Genetically Based Enzyme Deficiencies in
Humans
• Single gene mutations are responsible for many
human genetic diseases. Some mutations create a
simple phenotype, while others are pleiotropic
(affect many traits) (Table 4.2).
The lack of the enzyme that
transforms Phenylalanine into
tyrosine produces PKU.
The lack of the enzyme that
transforms the tyrosine into
DOPA, a precursor of
melanine produces albinism.
Phenylketonuria (PKU) is commonly caused by a mutation on
chromosome 12 in the phenylalanine hydrolase gene,
preventing the conversion of phenylalanine into tyrosine
(Figure 4.1).
Phenylalanine is an essential amino acid, but in excess it is
harmful, and so it is normally converted to tyrosine. Excess
phenylalanine affects the CNS, causing mental retardation,
slow growth, and early death.
PKU’s effect is pleiotropic. Some symptoms result from
excess phenylalanine. Others result from inability to make
tyrosine; these include fair skin and blue eyes (even with
brown-eye genes) and low adrenaline levels.
Albino British Blackbird. Photographed at the Natural
History Museum, South Kensington, London.
Adelie Penguin at Cape Bird
Antarctica
Name: Copito de Nieve / Floquet de Neu / Snowflake / Nfumu Ngui
Special characteristics: he was the first known albino gorilla
Albinism
•
Classic albinism results from an autosomal recessive
mutation in the gene for tyrosinase. Tyrosinase is used
to convert 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.
This enzyme defficiency is also found in a large
number of animals.
•
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.
Gene Control of Protein Structure
• Genes also make proteins that are not
enzymes. Structural proteins, such as
hemoglobin, are often abundant, making
them easier to isolate and purify.
• Sickle cell anemia is an example of how
genes control protein structure.
• J. Herrick (1910) first described sickle-cell
anemia, finding that red blood cells (RBCs)
change shape (form a sickle) under low O2
tension . Sickled RBCs are fragile and less
flexible than normal, hence the anemia and
capillaries blockage.
• Effects are pleiotropic, including damage to
extremities, heart, lungs, brain, kidneys, GI tract,
muscles, and joints. Results include heart failure,
pneumonia, paralysis, kidney failure, abdominal pain,
and rheumatism.
• The genes bA and bS are codominant.
Hemoglobin of bA/bA individuals has normal b
subunits, while hemoglobin of those with the
genotype bS/bS has b subunits that sickle at low
O2 tension. Hemoglobin of bA/bS individuals
is 1⁄2 normal, and 1⁄2 sickling form. These
heterozygotes may experience sickle-cell
symptoms after a sharp drop in the oxygen
content of their environment.
• The genes bA and bS are codominant.
Hemoglobin of bA/bA individuals has normal b
subunits, while hemoglobin of those with the
genotype bS/bS has b subunits that sickle at low
O2 tension. Hemoglobin of bA/bS individuals
is 1⁄2 normal, and 1⁄2 sickling form. These
heterozygotes may experience sickle-cell
symptoms after a sharp drop in the oxygen
content of their environment.