Information Restructuring

8/7/2019 Information Restructuring

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Processes involved in information

restructuring Restriction and modification

DNA repair mechanisms

Recombination Gene rearrangements

Gene amplification


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DNA METHYLATION

Occurs at the polynucleotide level, with

transfer of a methyl group from S-

adenosylmethionine

In eukaryotes, 5-methylcytosine is the

only methylated base

In prokaryotes, the major methylated

base is N6 methyl adenine Methylation in bacteria occurs at specific

sites


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Methylated bases


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DNA METHYLATION

In E.coli, methylation of A residues in the

sequence 5-----GATC-----3 is involved

in mismatch repair and it plays a central

role in controlling initiation of DNA

replication

Methylation at other sites protects DNA

cleavage by restriction enzymes


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DNA METHYLATION

In animals, methylation is foundprimarily in C residues that areimmediately 5 to G residues

Corresponding C in the complementarystrand is also methyated

In plants, the sequence is 5---CpNp--3

Methylation at a particular site isheritable (maintainance methylase)

Methylation occurs after replication


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DNA METHYLATION

Responsible for tissue-specific

inactivation of genes during development

Methylation is correlated with the arrestof expression of development-related

genes by altering the structure of

chromatin

Methylation may be involved incarcinogenesis (transition mutation)


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RESTRICTION and

MOD

IFICATIO

N

Restriction endonucleases

Nongenetic phenomena

Widespread among bacteria Restriction maps

Three types of restriction-modification

systems

Each type consists of a DNA methylase

and an endonuclease


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TYPE I

Have both enzymes in one protein

molecule

The protein molecule contains 3

subunits: nuclease, methylase,

sequence recognistion determinant


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TYPE II

Widely used in research because most

cut within the recognition sequence

Homodimers

Requires divalent cation for cleavage

but not ATP

A hemimethylated DNA is a preferred

substrate for methylase but nucleasecleaves only when the recognition sites

is unmethylated on both strands


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TYPE II

Cleavage generates a 3 hydroxyl and a

5 phosphate

Some enzymes generate a 5 over hang,

some 3 and others generate a blunt end


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TYPE III

Resemble type I

Contain both nuclease and methylase

activities in a 2 subunit enzyme

Do not require ATP

Modify just 1 strand of DNA

Cleavage site is fairly close to

recognition site


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DNAREPAIR

Defense of an organism to maintain

metabolic stability

Oxidative damage with concommitant

release of reactive oxygen species

causes the formation of DNA bases

such as 8-oxoguanine

Formation of this base contributes mostsignificantly to mutagenesis


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DNAREPAIR SYSTEMS

Direct repair

Nucleotide excision repair

Base excision repair Recombinational repair

Mismatch repair


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DNA

PHOTOPRODUCTS


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DIRECT REPAIR

PHOTOREACTIVATION

- DNA photolyase repair cyclobutanepyrimidine dimers

- 370 nm is the most effective wavelength

-the enzyme contains 2 chromphores:FADH- and 5,10-methenyltetrahydrofolateor 8-hydroxy-5-deazaflavin

-recent evidence suggests that humancells do not contain the enzyme


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DIRECT REPAIR

O6 AlkylguanineAlkyltransferase

-methylated or ethylated DNA iscomparable to UV irradiation because of

base modification-alkylation can block replication. Hence,some alkylating agents are used incancer therapy

-MNU, EMS, and MNNG are thecommon alkylating agents used in thelaboratory


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DIRECT REPAIR

O6 AlkylguanineAlkyltransferase

- bases altered are mostly purines

-O6

alkylguanine is the most mutagenicbecause of its high probability in pairing

with thymine thus, producing transition.

-the enzyme is not recyclable


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M

is

m

a

t

ch

R

ep

a

i

r


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RECOMBINATION

Maintains diversity

Any process that involves formation of

new DNA from distinct DNA molecules,

such that genetic information from each

parental DNA molecule is present in the

new molecules

4 types: homologous, site-specific,transposition and illegitimate


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HOMOLOGOUS RECOMBINATION

Recombinatorial events like meiotic

recombination, conjugation during

bacterial mating, bacterial

transformation and transduction allutilize homologous recombination

Most common way is to break and rejoin

DNA molecules


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SITE-SPECIFIC RECOMBINATION

Involves limited sequence homology

between recombining partners

Sites of breaking and joining are

determined by specific DNA-protein

interactions


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Characteristics ofDifferent Types

ofRecombination

TYPE

REQUIREMENTS

SEQUENCE

HOMOLOGY

RecA protein or

Counterpart

Sequence

specific enzyme

Homologous Yes yes No

Site-specific Yes (~ 15 bases) No Yes

Transposition No No Yes

Illegitimate no No Unknown


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GENE REARRAGEMENTS

Confers genome plasticity

Antibody diversity

Transposable elements Retroviruses


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ANTIBODYDIVERSITY


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ANTIBODYDIVERSITY


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TRANSPOSABLE ELEMENTS

Depending on whether the sequences

are oriented identically or in reverse,

homologous recombination between

them can yield deletions or inversions

Can activate or inactivate the gene into

which they move

Three classes of transposable elementsin bacteria depends on the involvement

of the transposase and resolvase


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CLASS I

Encodes a transposase but not a resolvase

Two types of class I: insertion sequenceand composite transposon

Insertion sequence is the simplesttransposable element.

It consists a gene for trasnposase flankedby 2 short inverted sequence repeats about15-25 bases pairs

Composite transposons consits of a proteincoding gene (antibiotic resistance) flankedby 2 insertion sequences


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CLASS II

Contain one set of short flanking direct

repeats

In addition to a protein-coding gene and

a transposase gene, a resolvase gene is

also present


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CLASS III

Belong to small group of bacteriophages

Phage Mu is the best studied

Inserts its chromosome at random in thehost chromosome by transpositional

mechanism and also replicate its

genome by the same mechanism

One gene encodes the transposase andanother gene encodes a protein with

DNA dependent ATPase activity


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Structures ofDifferent Classes of

Transposons


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TRANSPOSONS

Transposons and IS inserts at specific

traget sequence of 5 or 9 bp

Insertion involves duplication of that site

The two copies are present on each side

of the integrated element (action of

transposase)


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Information Restructuring

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