DNA repair, DNA Mutation, Gene Expression by Dr. Anurag Yadav

Various causes of DNA damage,
Methods of DNA repair for the Damage to the DNA structure,
Gene regulation and Gene Expression in eukaryotes and Prokaryotes.

1. Molecular Biology –DNA Repair,
Gene Mutation, Gene Expression
Dr Anurag Yadav
MBBS, MD
Assistant Professor
Department of Biochemistry
Instagram page –biochem365
YouTube –Dr Biochem365
Email: [email protected]
MNR MEDICAL COLLEGE & HOSPITAL

2. Content
DNA Damage
DNA repair Mechanism
Mutation
Gene Expression

4. DNA Damage & Repair
•Beingthecarrierofgeneticinformation,thecellular
DNAmustbereplicated(duplicated),maintained,and
passeddowntothedaughtercellsaccurately.
•Ingeneral,theaccuracyofreplicationisextremelyhigh.
•However,theredooccurreplicationerrors.
•Thecellsdopossesthecapabilitytorepairdamages
donetoDNAtoalargeextent.

5. Causes of DNA Damage
•Misincorporationof deoxynucleotidesduring
replication
•By spontaneous deaminationof bases during normal
genetic functions
•From x-radiation that cause “nicks” in the DNA
•From UV irradiation that causes thymine dimer
formation
•From various chemicals that interact with DNA.

7. MUTATIONS
Mutation: Change in the DNA structure of a
gene.
Mutagens: are Substance which bring about
(induce) mutation are collectively.
Changes that occur in DNA on mutation are
reflected in replication, transcription and
translation.

8. Types of Mutation
Two major
types
Point
mutation
Frameshift
mutation

9. 1. Point Mutation
•The replacement of one base pair by another
results in point mutation.
–Transition: a purine (or pyrimidine) is replaced by
another.
–Transversion: characterised by replacement by a
purine by a pyrimidineor vice versa.

10. 2. Frameshift Mutation
•Occurwhenoneormorebasepairsare
insertedinordeletedfromtheDNA.
•Causinginsertionordeletionmutation.

11. Consequences of Point Mutation
1.Silent Mutation: There is no detectable
effects.
Eg: UCA code is changed to UCU, but still codes
for Serine.
This is due to degeneracy of the genetic code.

12. Consequences of Point Mutation
2.Missensemutation:thechangedbasemaycode
forandifferentaminoacid.
Eg:UCAcodesforserine,replacedtoACAwhich
codesforthreonine.
Aminoacidmaybeacceptable,orPartially
acceptableorUnacceptable.
Eg:sicklecellanemia.

13. Consequences of Point Mutation
3.Nonsensemutation:sometime,thecodewith
thealteredbasemaybecomeatermination
codon.
Eg:secondbasechangeinUCAofserinetoUAA-
whichisaterminationcodon.
Causingprematureterminationoftheprotein
synthesis.

14. Consequences of Frameshift
mutations
•Theinsertionordeletionofabaseinagene
resultsinanalteredreadingframeofthemRNA.
•Thetranslationcontinueswithnewcodon
inserted.
•Resultinproteinwithseveralalteredaminoacids
orprematureterminationofproteinsynthesis.

15. REPAIR OF DNA
Cell possess the inbuilt system to repair
the damaged DNA.
•Base excision repair
•Nucleotide excision repair
•Mismatch repair
•Doubled strand break repair
Achieved by
4 distinct
mechanisms.

16. 1. BASE EXCISION REPAIR
•Thebasescytosine,adenineandguaninecan
undergospontaneousdepurinationto
respectivelyformuracil,hypoxanthineand
xanthine.
•Thesealteredbasesdonotexistinthenormal
DNA,andthereforeneedtoberemoved.
•Thisiscarriedoutbybaseexcisionrepair

18. 2. NUCLEOTIDE EXCISION REPAIR
•TheDNAdamageduetoultravioletlight,ionizing
radiationandotherenvironmentalfactorsoften
resultsinthemodificationofcertainbases,
strandbreaks,cross-linkagesetc.
•Nucleotideexcision-repairisideallysuitedfor
suchlarge-scaledefectsinDNA.

19. 2. NUCLEOTIDE EXCISION REPAIR
•Aftertheidentificationofthedefectivepieceofthe
DNA,theDNAdoublehelixisunwoundtoexpose
thedamagedpart.
•Anexcisionnuclease(exinuclease)cutstheDNAon
eitherside(upstreamanddownstream)ofthe
damagedDNA.Thisdefectivepieceisdegraded.
•Thegapcreatedbythenucleotideexcisionisfilledup
byDNApolymerasewhichgetsligatedbyDNAligase

21. •Xerodermapigmentosum(XP)isarare
autosomalrecessivedisease.Theaffected
patientsarephotosensitiveandsusceptibleto
skincancers.ItisnowrecognizedthatXPis
duetoadefectinthenucleotideexcision
repairofthedamagedDNA.

22. 3. MISMATCH REPAIR
•Despitehighaccuracyinreplication,defects
dooccurwhentheDNAiscopied.
•Cytosine(insteadofthymine)couldbe
incorporatedoppositetoadenine.
•Mismatchrepaircorrectsasinglemismatch
basepaire.g.CtoA,insteadofTtoA.

23. Template strand
of the DNA
exists in a
methylated
form,
while the newly
synthesized
strand is not
methylated.

24. •Hereditarynonpolyposiscoloncancer
(HNPCC)isoneofthemostcommon
inheritedcancers.Thiscancerisnowlinked
withfaultymismatchrepairofdefectiveDNA.

25. 4. DOUBLE-STRAND BREAK REPAIR
•Double-strandbreaks(DSBs)inDNAare
dangerous.
•Theyresultingeneticrecombinationwhichmay
leadtochromosomaltranslocation,broken
chromosomes,andfinallycelldeath.
•DSBscanberepairedbyhomologous
recombinationornon-homologousendjoining

28. GENE EXPRESSION AND REGULATION
•Organisms adapt to environmental changes by
altering gene expression.
•The regulation of the expression of genes is
necessary for the growth, development,
differentiation and the very existence of the
organism.

29. TYPES OF GENE EXPRESSION
TWO MAIN
TYPES
POSITIVE
REGULATION
NEGATIVE
REGULATION

30. 1. POSITIVE REGULATION
•Whentheexpressionofgeneticinformationis
quantitativelyincreasedbythepresenceof
specificregulatoryelement,itiscalledas
positiveregulation.
•Theelementormoleculemediatingpositive
regulationiscalledpositiveregulator.

31. 2. NEGATIVE REGULATION
•Whentheexpressionofgeneticinformationis
decreasedbythepresenceofaspecific
regulatoryelement,itiscalledasnegative
regulation.
•Theelementormoleculemediatingthenegative
regulationiscalledanegativeregulator.

32. ONE CISTRON-ONE SUBUNIT
CONCEPT
•Earlierhypothesisproposedthatonegene
producesoneenzymeorproteinand“onegene-
oneenzyme”conceptwasintroduced.
•Itisnowknownthatsomeenzymesandprotein
moleculesarecomposedoftwoormore
nonidenticalsubunits,whichcannotbeexplained
by“onegene-oneenzyme”theoryandsoitisnot
valid.

33. •Thecistronisnowconsideredasthegeneticunit
codingforthestructureofthesubunitofan
enzymeorproteinmolecule,actingasitdoesas
thesmallestunitofgeneticexpression.
•Hence,the“onegene-oneenzyme”ideamight
bemoreaccuratelyregardedas“onecistron-one
subunitconcept”.

34. THE OPERON CONCEPT
•TheOperonisthecoordinatedunitofgenetic
expressioninbacteria.
•TheconceptofOperonwasintroducedby
JacobandMonodin1961.
•regulationoflactosemetabolisminE.coli.
ThisispopularlyknownasLACOPERON.

35. Operon is defined as a segment
of a DNA strand consisting of:
•Structuralgenes
•Operatorgene
•Regulatorgene
•Psite(Promotersite)

36. Lac-operonStructure
consists of a regulatory gene (I; I for inhibition),
operator gene (O) and
three structural genes (Z, Y, A).
Besides these genes, there is a promoter site (P), next to the
operator gene, where the enzyme RNA polymerase binds.
The structural genes Z, Y and A respectively, code for the
enzymes β-galactosidase, galactoside permease and galactoside
acetylase.

37. •β-Galactosidasehydrolyseslactose(β-
galactoside)togalactoseandglucosewhile
permeaseisresponsibleforthetransportof
lactoseintothecell.

39. Repression of Lac operon
•Theregulatorygene(I)isconstitutive.Itis
expressedataconstantrateleadingtothe
synthesisoflacrepressor.
•lacrepressorbindstooperatorgene(O).
•PreventsbindingofenzymeRNApolymeraseto
promotersite(P).
•TherebyblockingthetranscriptionofStructural
gene.

40. DEREPRESSION OF LAC OPERON
•Inthepresenceoflactose(inducer)inthe
medium,asmallamountofitcanentertheE.
colicells.Therepressormoleculeshaveahigh
affinityforlactose.
•Thelactosemoleculesbindandinducea
conformationalchangeintherepressor.

41. •Repressorgetsinactivatedandcannotbindto
operatorgene(O).
•RNApolymeraseattachestoDNAatpromoter
siteandtranscriptionproceeds.
•LeadingtoformationofpolycistronicmRNA
andfinally3enzymesareformed.
•Thuslactoseinducesynthesisof3enzymes.

44. GENE EXPRESSION IN EUKARYOTES
•Eachcellofthehigherorganismcontainsthe
entiregenome.Asinprokaryotes,gene
expressionineukaryotesisregulatedto
providetheappropriateresponsetobiological
needs.
•Thismayoccurinthefollowingways

45. •Expressionofcertaingenes(housekeeping
genes)inmostofthecells.
•lActivationofselectedgenesupondemand.
•lPermanentinactivationofseveralgenesinall
butafewtypes

46. •Histoneacetylationanddeacetylation
•MethylationofDNAandinactivationofgenes

48. Important Q
•Mutation
•Types of Mutation
•Clinical consequence of Mutation
•Causes of DNA damage
•DNA repair mechanisms
•Gene Expression and Regulation
•Types of Gene regulation