DNA repair and recombination

DNA
INTRODUCTION
CHEMICAL COMPOSITION
NUCLEOSIDES & NUCLEOTIDES
DNA REPAIR
INTRODUCTION
TYPES OF DNA REPAIR
I)DIRECT REPAIR SYSTEM,
II)BASE EXCISION REPAIR,
III)NUCLEOTIDE EXCISION REPAIR,
IV)MISMATCH REPAIR,
V)RECOMBINATION REPAIR,
DEFECTS IN DNA REPAIR UNDERLIE HUMAN DISEASE
DNA RECOMBINATION
INTRODUCTION
MECHANISM OF DNA RECOMBINATION
TYPES OF RECOMBINATION
I) HOMOLOGOUS RECOMBINATION
MODELS FOR HOMOLOGOUS RECOMBINATION:-
I)HOLLIDAY MODEL,
II)MESSELSON AND RADDING MODEL,
III)DOUBLE STRAND BREAK MODEL,
GENE CONVERSION
II) NON-HOMOLOGOUS RECOMBINATION,
i) SITE SPECIFIC RECOMBINATION,
ii)TRANSPOSITIONAL RECOMBINATION.,

1. DNA REPAIR AND RECOMBINATION
By
KAUSHAL KUMAR SAHU
Assistant Professor (Ad Hoc)
Department of Biotechnology
Govt. Digvijay Autonomous P. G. College
Raj-Nandgaon ( C. G. )

2. SYNOPSIS:
DNA
INTRODUCTION
CHEMICAL COMPOSITION
NUCLEOSIDES & NUCLEOTIDES
DNA REPAIR
INTRODUCTION
TYPES OF DNA REPAIR
I)DIRECT REPAIR SYSTEM
II)BASE EXCISION REPAIR
III)NUCLEOTIDE EXCISION REPAIR
IV)MISMATCH REPAIR
V)RECOMBINATION REPAIR
DEFECTS IN DNA REPAIR UNDERLIE HUMAN DISEASE

3. DNA RECOMBINATION
INTRODUCTION
MECHANISM OF DNA RECOMBINATION
TYPES OF RECOMBINATION
I) HOMOLOGOUS RECOMBINATION
MODELS FOR HOMOLOGOUS RECOMBINATION: -
I)HOLLIDAY MODEL
II)MESSELSON AND RADDING MODEL
III)DOUBLE STRAND BREAK MODEL
GENE CONVERSION
II) NON-HOMOLOGOUS RECOMBINATION
i) SITE SPECIFIC RECOMBINATION
ii)TRANSPOSITIONAL RECOMBINATION.

4. DEOXYRIBOSE NUCLEICACID(DNA):-
•DNAispresentincellsofallplants,animals,prokaryotesandano.
ofviruses.
•Ineukaryotes,itispresentinsidethenucleus,chloroplastand
mitochondriawhereasinprokaryotes,itisdispersedincytoplasm.
•TheDNAofallplants,animals,&manyvirusesisdoublestranded
exceptsomeviruses.Eg.Psix174.
•InsomevirusesthegeneticmaterialisRNA.

5. CHEMICAL COMPOSITION OF DNA: -
•DNA is a polymer of deoxyribonucleotide monomer.
•DNA is formed of three important component i.e.
•Pentose Sugar:-In DNA deoxyribose is a pentose sugar.
•InDNA,allthesugarsinonestrand
aredirectedtooneendi.e.strandhas
polarityandsugarsoftwostrandsare
directedinoppositedirection.

6. •Nitrogenousbases:-
Twotypes
•Purine:-Theyaredouble
ringcompounds.InDNA
twopurinesareAdenine(A)
andGuanine(G).
•Pyrimidine:-Theyaresingle
ringcompounds.InDNA
twoPyrimidinesareCytosine
(C)andThymine(T).
•PhosphateGroup:-ItContainsalpha,beta,gammaphosphate.
•IntheDNAstrand,thephosphategroupalternatewith
deoxyribose.

7. NucleosidesandNucleotides:-
NUCLEOSIDE=Sugar(deoxyribose)+Nitrogenousbases.
NUCLEOTIDE =Nucleoside+phosphategroup.
•Inotherwords,Anucleosideisabase-sugarcombinationanda
nucleotideisanucleosidephosphate.
•In1953,WatsonandCrick,proposedthedoublehelicalmodelof
DNAandisthemostwidelyacceptedmolecularmodelofDNA.

8. Fig:-DNA Double Helix.

9. DNA REPAIR:-
•DNAhasaspecificbasesequenceandthemainobjectiveof
biologicalsystemistomaintainthesesequencesofDNAfromone
generationtoother.
•Inamulticellularorganism,thedeathofasinglesomaticcelldueto
DNAdamageislessdangerousthantoreplicateitsdamagedDNA
,asitgiverisetotumourorothercancerousgrowth.
•Thus,cellmustpossessanefficientrepairsystemtoavoidcelldeath
ormutation.
•Thisrepairsystemremovesmanyspontaneouserrorsthatarisesdue
tochemicalmutagensandradiationsandalsomanydamagesarose
duringreplication.

10. Types of DNA repair:-
•MostcellspossessfourdifferentcategoriesofDNA
repairsystem.(LindahlandWood,1999):

11. •DIRECT REPAIR SYSTEM: -
•NicksarerepairedbyaDNAligase.Nicksoftenresultsfromthe
effectsofionizingradiation.
Fig; Repair of a nick by DNA Ligase.
•Onlyafewtypesofdamagednucleotidecanberepaireddirectly:i.e.

12. •Someformsofalkylationdamagearedirectlyreversibleby
enzymesthattransferthealkylgroupfromthenucleotidetotheir
ownpolypeptidechains.
Eg.AdaenzymeofE.coli,itisinvolvedinanadaptiveprocessthat
thisbacteriumisabletoactivateinresponsetoDNAdamage.Ada
removesalkylgroupandcanalsorepairphosphodiesterbondsthat
becamemethylated.
•Cyclobutyldimersarerepairedbyalight-dependentdirectsystem
calledphotoreactivationInE.coli,theprocessinvolvestheenzyme
calledDNAphotolyase.Whenstimulatedbylight(wavelength
between300and500nm)theenzymebindstocyclobutyldimers
andconvertsitbacktotheoriginalmonomericnucleotides.
Asimilartypeofphotoreactivationinvolvesthe
(6-4)photoproductphotolyaseandresultsinrepairof(6-4)lesions.

13. •EXCISION REPAIR:-
•These pathway is divided into two categories:
•Base Excision Repair
•Nucleotide Excision Repair
•BaseExcisionRepair:-
•Itinvolvesremovalofadamagednucleotidebase,&excisionofa
shortpieceofthepolynucleotidearoundtheAPsitecreated,and
resynthesiswithaDNApolymerase.

14. Fig:Base excision repair

15. NUCLEOTIDE EXCISION REPAIR:-
•Nucleotideexcisionrepairhasamuchbroaderspecificitythan
baseexcisionsystem.
•Usedtocorrectmoreextensivetypeofdamage.
•Eg.intrastrandcrosslinks&modifiedbasesbychemical
mutagens.
•similartobaseexcisionexceptthatitisnotprecededbyselective
baseremoval,andalongerstretchofpolynucleotideisexcised.
•RepairmaybeShortpatchorLongpatch.
•Italsocorrectcyclobutyldimersbyadarkrepairprocess(for
thoseorganismthatdonothavephotoreactivationsystem).

16. Fig; Short patch nucleotide-excision repair in E.coli.
•Shortpatchprocessofnucleotideexcisionrepairis
generallystudiedbyeg.ofE.coli.

17. Fig:-Nucleotide excision repair in eukaryotes.
•E.colialsohasalongpatchnucleotideexcisionrepairthatinvolves
excisionofDNAupto2kbinlengthandtheeukaryoticnucleotide
excisionrepairprocessisalsocalled‘longpatch'repair.

18. MISMATCH REPAIR SYSTEM: -
•Itcorrectsthemismatchesresultingfromerrorsin
replication.
•Onceamismatchisfound,therepairsystemexcisespart
ofthedaughterpolynucleotideandfillsinthegap,ina
mannersimilartobaseandnucleotideexcisionrepair.
•E.colihasthreemismatchrepairsystems,called‘long
patch',‘shortpatchand‘veryshortpatch',

19. Fig. Long patch mismatch repair in E. coli.
•ThelongpatchsystemrequirestheMutH,MutLandMutSproteins,as
wellastheDNAhelicaseIIduringmismatchrepair.
•Similareventsoccurduringshortandveryshortmismatchrepair,
thedifferenceisthespecificitiesoftheproteinsthatrecognizethe
mismatch.

20. RECOMBINATION REPAIR: -
•Recombination repair is used to mend double-strand breaks.
•Doublestrandbreakrepairprocess
involvesfoursetsofgenes(Critchlow
andJackson,1998).Thesegenes
specifyamulti-componentprotein
complexthatdirectsaDNAligaseto
thebreak.
•Thecomplexincludesaprotein
calledKu,whichbindstheDNAends
eithersideofthebreak.
Fig. Single-and double-strand-break repair.

21. •RecombinationrepairprocessisalsocalledNon-
homologousEndJoining(NHEJ),indicatingthatthereis
noneedofhomologybetweentwomoleculeswhoseend
arebeingjoined.
•NHEJisalsoatypeofrecombination,asitisusedtojoin
togethermoleculesthatwerenotpreviouslyjoined,
producingnewcombinations.
•AversionoftheNHEJsystemisprobablyusedduring
constructionofimmunoglobulinandT-cellreceptor
genes,butthedetailsarelikelytobedifferent.

22. •DNArepairisveryimportantasitcanbeanalysedbytheno.and
severityofinheritedhumandiseasesthatarelinkedwithdefectsin
oneoftherepairprocesses.
•Foreg:Xerodermapigmentosum,itiscausedduetomutationinone
geneforproteinthatisinvolvedinnucleotideexcisionrepair(NER).
•TRichothiodistrophyisalsocausedbydefectsinNER,butitismore
complexasitincludeproblemswithboththeskinandthenervous
system.
•Afewdiseasesarelinkedwithdefectsinthetranscriptioncoupled
componentofnucleotideexcisionrepair.
RESULTSOFDEFECTSINDNAREPAIR:-

23. DNA RECOMBINATION
•GeneticrecombinationisaprocessbywhichasegmentofDNAis
brokenandthenjoinedtoadifferentDNAmolecule.
•Recombinationisofgreatevolutionarysignificance,asitbrings
aboutvariation.
•Thus,recombinationprovidesameansforspreadingoffavourable
allelesandameanstoeliminateanunfavourableallele.
•Recombinationwasfirstrecognizedastheprocessresponsiblefor
crossing-overandexchangeofDNAsegmentsbetween
homologouschromosomesduringmeiosisofeukaryoticcells,and
wassubsequentlyimplicatedintheintegrationoftransferredDNA
intobacterialgenomesafterconjugation,transductionor
transformation.
INTRODUCTION:-

24. •MechanismoFRecombination:-Basically,therearethreetheories-
•Breakageandreunion:-Twohomologousduplexofchromosome
lyinginpairedformbreaksandrejoincrosswiseandrecombinants.
•Breakageandcopying:-Onehelixofpairedhomologous
chromosomebreaksintwosegmentsandoneofthemisreplacedby
anewlysynthesizedsegmentcopiedfromotherhelixofpaired
chromosome.
•Completecopychoice:-(Belling,1931)Accordingtothistheory,a
portionofoneparentalstrandofhomologouschromosomeactsas
templateforsynthesisofcopyofitsDNAmolecule.Theprocessof
copyingshiftstootherparentalstrand.
Thus,recombinantscontainsomegenetic
informationofoneparentalstrandandsomeoftheotherstrand.

25. A.)Breakage and
reunion
B.)Breakage
and copying
C.)Complete copy
choice
Fig.Three possible Mechanism of recombination
a b a
b+
a+ b+
a+b

26. TYPES OF RECOMBINATION: -
Recombination is divided mainly into two categories:-
•Homologous recombination
•Non-Homologous recombination
HOMOLOGOUS RECOMBINATION:-
•Homologous recombination(alsocalledGeneralized
Recombination)involvesexchangeofpreciselycorresponding
sequencesbetweenhomologousDNAduplexes.
•ItisareactionbetweentwoduplexesofDNA,andtheenzyme
responsibleforthiscanuseanypairofhomologoussequencesas
substrates.

27. Models For Homologous Recombination:-
I.The Holliday model :-
•ProposedbyRobinHollidayin
1964.
•Re-establishedbyDavidDressler
andHuntingtonPotterin1976,
whodemonstratedthatthe
proposedphysicalintermediates
existed
Fig; Holliday model for the homologous recombination.

28. II.MeselsonandRaddingModel:-
•Meselson and Radding in 1975,
proposes a model, and is a more
satisfactory scheme.
Here, a single stranded nick occurs
in just one of double helices
Fig: Meselson & Radding model.

29. III.Doublestrandbreakmodel:-
•Accordingtothismodel,adouble
strandbreakinoneDNAmolecule
initiatestheprocessofrecombination.
Fig: Double strand break model.

30. Fig:Resolution of two holliday junction in double strand break model.
Double strand break recombination results in two holliday
junction , which can be resolved in two ways:-

31. GENE CONVERSION: -
•GeneconversionisaneventinDNAgeneticrecombination,it
occursathighfrequenciesduringmeioticdivisionbutalsooccursin
somaticcells.
•Whenapairofallelesegregates,tetradsareexpectedtohave2:2
ratioofsporesbutsometimethisexpectedpatternisreplacedby
3:1ratio.ThisiscalledGeneconversion.
•ItisaprocessbywhichDNAsequenceinformationistransferred
fromoneDNAhelix(whichremainsunchanged)toanotherDNA
helix,whosesequenceisaltered.
•Geneconversionhasoftenbeenrecordedinfungalcrosses.

32. Fig: Gene conversion.

33. NON-HOMOLOGOUS RECOMBINATION:-
•Alsocalledasspecializedrecombination.
TYPES OF NON-HOMOLOGOUS RECOMBINATION:-
•SITE-SPECIFIC RECOMBINATION
•TRANSPOSITION

34. •SITE-SPECIFIC RECOMBINATION: -
•Site-specificrecombinationinvolvesareactionbetweentwo
specificsites.
•Here,aregionofextensivehomologyisnotrequired,the
processcanalsobeinitiatedbetween2DNAmoleculethathave
veryshortsequencesincommon.
The classic model for site-specific
recombination is illustrated by phage
lambda:

35. TRANSPOSITIONAL RECOMBINATION:
•Transpositionis not a type of recombination but a process that
utilizes recombination, the end result being the transfer of a
segment of DNA from one position in the genome to another.
•Transposable element could be broadly divided into three
categories on the basis of their transposition mechanism:-
•DNA transposonsthat transpose replicatively, the original
transposon remaining in place and a new copy appearing elsewhere
in the genome.
•DNA transposonsthat transpose conservatively, the original
transposon moving to a new site by a cut-and-paste process.
•Retroelements, all of which transpose via an RNA intermediate.

36. ReplicativeAndConservative
Trasnsposition:-
•Manymodelshavebeen
proposedovertheyearsbut
mostaremodificationsofa
schemeoriginallyoutlinedby
Shapiro(1979).
A model for the process resulting in replicative and conservative transposition.

37. •TranspositionofRetroelements:-
•Ittakesplaceintwosteps:-
•1
st
step:-Anintegrated
retroelementiscopiedintoa
freedouble-strandedDNA
version.Firstly,thereis
synthesisofanRNAcopy,
whichisthenconvertedto
double-strandedDNAbya
seriesofeventsthatinvolves
twotemplateswitches.

38. ii) 2
nd
step:-is the transposition of the retroelement.
Fig: Transposition of a retroelement :Part II.

39. One of the basic feature of the organisms, are to adapt in the
environment, and DNA repair & recombination helps the
organism to adapt to this feature .
DNA repair help to correct the error of replication and also the
damage caused due to chemical mutagens and radiations.
And, DNA recombination brings about variation and thus, leads to
evolution.
Recombination helps to distinguish between the favorable and
un-favorable changes occurred due to mutation and forms the
basis for natural selection.
Thus, repair and recombination, both play
an important role in the biological system.
CONCLUSION:-

40. REFERENCES:-
Genetics B.D.Singh
Textbook of Biochemistry David M. thomsan
Genome T.A.Brown
Gene VIII
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