Download Molecular and Biochemical Basis of genetic Disorder

Molecular and Biochemical Basis
of
genetic Disorder

Prof. Dr. Müjgan CENGİZ
Principles of molecular disease
* Molecular reason of a genetic
disease is a mutation. This
mutation either inherited or
acquired.
*The biochemical genetic is study of
phenotype at the level of proteins,
biochemistry and metabolism.
A genetic disease occurs when an
alteration in the DNA of an essential
gene changes the amount or function
or both, of the gene product -mRNA
and protein.
 Single Gene disorders almost always
results from mutations that alter the
function of a protein.
 The pathologies because of the
defaults of a protein expression can

1-Mutations at the control genes.
 2-Mutations at the structural genes.
 The mutations at the control genes
causes single nucleotide change. The
amount of the polypeptide and
function does not effected.
 Only the velocity of protein synthesis
changes, phenotype is normal.

Mutations taking place at the structural
genes causes a genetic disease .
 The amounts and functions of gene
products decreases.
 To uderstand the pathogenesis of a
genetic disease we have to learn the
knowledge of primary abnormalities
the results of the alteration of the gene
function.

Disease causing mutations have been
identified in more than 1990 of the 3310
single gene disease.
 There are four effects of disease causing
mutations on protein function.
 1- Loss of function of a protein.
 2-Gain a function.
 3-Acquision of a novel property by mutant
protein.
 4- Expression of a gene at the wrong time
or place.

Examples of these effects:
1-α thalasemias are commonly examples for
deletions of α globin genes.
2-Gain function mutations can alter the
biochemical phenotype by increasing the
function of a protein.
This effect because of
a-İncrease in the level of protein’s
expression(Trisomy 21).
b- İncrease in the ability of each protein to
perform one or more normal function.
Achondroplasia (fibroblast growth factor
mutation active in the absence of the ligand).
3-Acquising of a novel property:In a
few disease the change in amino acid
sequence does not change function.
Ex.Sickle cell disease(β chain:Glu6Val)
no effect on on transport of oxygen but
this hemoglobin chains aggregate when
deoxigenated, it forms polymeric fibres.

4-Expression of a gene at the wrong
time: the regulatory regions of the
gene can alter and cause an
inappropriate expression of the gene
in normal time or place. ex oncogene
normally promotes cell proliferation.
But if is not normally expressed
causes cancer.
Some mutations in hemoglobin regulatory
elements lead to the continued expression
in the adult life γ globulin gene.
 This gama globulin is normally expressed
at high levels only in fetal not postnatal
life.
 This mutation led to phenotype called
persistance of fetal hemoglobin.

The molecular and Biochemical causes of
genetic diseaes:
 Mutation protein coding
genes(DNA)→mutation→Defective
protein production.
 These are examples of single gene
disorders. Some examples of proteins
associated with monogenic diseases.

Some examples of Classes of Proteins
Associated with Monogenic Diseases
1-Transport and storage
 İnterorgan Hemoglobin
(thalassemias)(AR)
 İntracellular transport (copper
transport prot. menkes syndr. (AR)
 Epitel membr. Cystic fibrosis (CFRT
AR)

2-Enzyme defects
 Amino acids - PKU(phenyl alanine hydroxilase
AR)
 Complex lipid-Tay sachs(Hexosaminidase A AR)
 Purines-immundeficiency( Adenosine deaminidase
AR)
 Carbohydrates Galactose 1 phosphate uridyl
transferase
3-Structure of cells and organs
 Duschene muscular distrophy(dystrophin XR)


4-Control of growth and differantiation-Tumor
suppressors,RB gene products (AR),
oncogenes(AD).
5-Intracellular metabolism and comunication
Growth gormon(dwarfizm,AR),insulin(AD)

Familial hypercholesterolemia(LDL receptor).


1-Transport and Storage
THALASSEMIAS
Hemoglobin is the oxygen carrier in
vertebrate red blood cells.
 The molecule contains four subunits
 2 α , 2 βglobin chains
 Each subunit composed of polypeptide
chain globin and prostetic group heme.
 Heme is a iron containing pigment that
combines with oxygen to give the
molecule its O transport ability.

Thalassemia is drived from greek
word of sea –Thalassa,miamediterenian origin.
 Hemoglobin A is normal adult
hemoglobin
 2 α 141 aa chain folded and fitted
 2 β 146 aa
 Four chain folded and fitted
 The chains resemble each other.

β thalassemias
 Decreasd β globin production causes
hypochromic, microcytic anemia.

Imbalance in globin synthesis leads
to precipitation of excess α chains,
leads damage of red cel membrane.

β chain is important in post natal
period.
Onset of β thalassemia
apperent untill a few monts after birth (β
globin replaces γ ).
 Synthesis of Hb A reduces.

Beta thalassemias usually single base
pair mutations.
 They are so many different type of
mutations.
 1- 2β thalassemia-allel thalassemia
major(severe anemia)
 2-So little production of βglobin No Hb A
present. β° thalassemia.
 3- Some Hb A is detectable. β+
thalassemia
 4-Carriers of one β+ thalassemia are
clinically thalassemia minor.

Thalassemia minor patients have
hypochromic,microcytic red blood cells
.
 They may have a slight anemia that
can miss diagnosed.

MOLECULAR BASIS OF β THALASSEMIA
Simple β thalassemias results many different types of
molecular abnormalities mostly point mutations in the β
globin gene.
 Deletions
-β globin gene deletions (619 base pair deletion)
 Defective mRNA synthesis
- RNA splicing defects
- promoter mutations
- poly adenylation signal defects
 Nonfunctional mRNA
- Nonsense mutations
- Frameshift mutations

TRANSPORT DEFECTS
CYSTİC FİBROSİS
 Autosomal recessive genetic disorders in Caucasian
population

İncidence ~ 1/ 2500

CF gene (called CFTR - Cystic fibrosis conductance
Regulatory protein)

The protein encoded by the CTFR gene is a regulated
Cl¯ channel located in the apical membrane of epithelia
cells affected by the disease.
PHENOTYPES
Lungs,
 Pancreas (deficiency of pancreatic enzymes)
 Sweat glands (eleveted level of Cl. >60meq/lt)
 Chronic obstructive lung disease ( because of thick
secretion and recurrent infection)
 Infertility (in male)
 Half of the patients survive to 26 years
 CF gene located on chorosome 7q31 spans about 250
kb DNA
 It has 27 exons
 Encode large integral protein 170 kD.
 Gene called CFTR

CTFR Cl¯ channel has 5 domain:
1. MSD1 (membrane spaining domain)
2. MSD2
3. NBD1 ( nucleotid binding domain) (AT
binding
domain)
4. NBD2
5. Regulatory domain

The pore of Cl channel is formed by the
12 transmembrane segments.
ATP is bound and hydrolyzed by the
NBDs.
Energy released is used for ion
transport.
Regulation of the channel is mediated by
phosphorylation of the R domain.
It has 4 type mutation
-class I : Defect in protein production
Premature stop codons or
mutations generates
-class II : Defective protein processing due
to misfolding of
protein
 Deletion at phenylalanine residue at
position 508 (∆F 508)
 The first ATP binding fold (NBD1)
 Accounts ~ 70% of CF allels

GER → Golgi →Plasma membrane
-class III : Mutations of the NBDs
Defective regulation of
protein
S1255P (mutation at NBD2)
-class IV : Mutation at regulatory
domain
Defective conduction due to
alteration of Cl¯ channel (R334W)

Detection and Treatment

Detection of Na and Cl amount

DNA mutation analysis

Prenatal Diagnosis

Population screening
2-ENZYME DEFECTS
A-AMINOACIDOPATHIES
Hyperphenylalainemias
 The abnormalities that lead to an increase in the
blood level of phenylalaline, is called PKU.
 There is mutations at phenylalaline coding gene
loci.
 There is loss of function mutations on the gene
encoding PAH.
 PAH gene isolated in 1986, more than 400 different
allels have been recognized.

Phenylalaninehydroxylase gene
mutations
Arg 408 trp- activity<1% -- %31
european
 Tyr414Cys- activity30% -- %5
“
 Ile 65 Thr -- activity 25% --%5
“

B-LYOSOSOMAL STORAGE DISEASES
Lyososomes are membrane bound organels
containing hydrolitic enzymes.
 They involved in the degradation of biologic
macromolecules.
 Genetic defects in these hydrolases lead to the
accumulation of their substrates in the lysosome.
 They lead cellular dysfunction-cell death .
 When brain is affected it causes
neurodegeneration.
 There are more than 48 lyososome hydrolase.
Almost all autosomal recessive.

TAY-SACHS DISEASE
Lyososomal storage disease (GM2
gangliosides)
 The inability to degrade a sphingolipid GM2
ganglioside.
 The biochemical lesion is deficiency of
hexoaminidase A.
 The disease has its clinical impact especially on
the brain.
 Hex A is the product of 3 gene system.
α subunit Hex A
β subunit Hex B genes
Mutations of the Hex A
Mutation
Effect
-4 bp insertion
premature
(exon 11)
-Exon 12 splice
Junction G→C
homozygote
phenotype
Tay-Sachs
stop codon
defective
mRNA spilicing
Tay-Sachs
3-DISORDER OF STRUCTURAL PROTEINS
DUCHENE-AND –BECKER MUSCULAR
DISTROPHY
 Severe untreatable, relatively common
X linked disorder.
 Related with Dystrophin protein.
 Affected boys normal at first year.
 Muscle weakness developed age 3-5.
 The child confined to wheelchair at age
12 and deads at the age of 20.
 Creatine kinase level is elevated 50100 times upper limit of normal
Patients die of respiratory failure or
cardiac failure, brain is also effected.
 Becker (BMP) is also due to mutation
in dystrophin gene
 İt produces a phenotype is much
milder.
 If patients is still walking at the age of
16 the disease is BMD.

İncidence of DMDis 1 in 3300 live male
birth
 Calculated mutation rate is (10)-4
 Normal male produces sperm with new
mutation of DMD gene every 10 to 11
seckonds(mens produces 8.107
sperm/day)
 Carier females have no clinical
manifestations. 70% have↑creatine
kinase
 İn rare instances females have been
reported with DMD.

Size of DMD gene 2300 kb
 %1.5 of X chromosome
 Gene is a complex gene it have 79
exons 7 tissue spesific promoter.
 It encodes 427 kD protein.

Mechanism of mutation in Duchenne
or Becker muscular distrophy
 Gene deletion
%60 dmd or bmd
 Point mut
%34
"
 Partial duplication %6
"
