mol bio unit 3

how do nucleotides link —> nucleic acids

3’OH attacks alpha phosphate of new nucleotide —> releases PPi and extends nucleotide via phosphodiester bond formation

RNA structure explains reactivity

existence of 2’OH allows lariat formation

backbones may spontaneously break

b form double helix

constant diameter

a form

dehydrated

z form

in vitro formation

minor vs major groove

diff amts of h bond donors + acceptors

seq- spec binding in major groove bc higher var of H bond donors + acceptors

non seq-spec in minor groove

DNA replication

one dsDNA —> 2 identical copies

DNA synth moves toward 5’ end of template on each strand

ori

used only once per cell cycle during S phase

have nearby GC righ regions

initiation of DNA replication

5’ end initiated by DNA primase

DNA pol can’t begin synth w/o existing 3’ end but primase can start anywhere

dna pol palm

active site w/ 2 atoms (usually Mg2+) allow phosphate groups’ charges to come tg

how DNA pol selects the right dNTP

1. finger and palm fit perf around correct dNTP

2. recognizes correct BP through minor groove by forming H bonds w it

3. proofreading via wrist

   1. incorrectly paired base not stable in active site

   2. wrong nucleoside enters editing site where backbone is cut by 3’ —> 5’ exonuclease (last nucleoside suffers same fate)

nucleosome re-establishment during replication

1. increase in histone synth in G1/S phase —> histone mod occur BEFORE replication fork is made

2. replisome isn’t blocked by histones

3. wave of H3k9ac precedes replication form

4. H3/H4 tetramers stay assoc w one strand or the other but h2a/h2b unbind + rebind

5. chr remodelling complexes aid with histone displacement

end replication problem (not in bacteria)

start with template strand with 3’ end

• when primer removed —> 3’ overhang

solution to end replication problem

telomerase complex binds close to 3’ end of overhanging template strand

adds dNTPs to extend DNA strand using a bound RNA template seq

repeats many times to build repetitive telomere seq

3’ end becomes long

primer added near 3’ end of longer strand —> dna pol + ligase

DNA replication summary

1. opening/ separation of dna double helix at origin of replication

2. topiso in front of helicase to relieve supercoiling

3. DNA + pcna clamp on both leading and lagging

   1. add dNTPs to 3’ end of primer strand

4. lagging strand has ssBPs

5. rna primers removed + replaced by DNA pol

6. ligase seals gaps

dna pol alpha

makes RNA primer

dna pol delta

makes lagging strand

dna pol epsilon

makes leading strand

RNAse

remove RNA primers

helicase

separates template strands

PCNA clamp

holds leading + lagging strand pol —> clamp loader helps PCNA

topoiso

cut DNA backbones to relieve overwinding/ supercoiling

tautomers

resemble normal bases

usually change back to normal and removed by proofreading before dna pol replicates more bases

if DNA pol moves on before removal —> mismatch

slippery seq

seq w multiple identical repeated seq —> strands can bind tg and loop —> indels when replicating and lead to 3, 4, 5 nt repeats

do somatic cells express telomerase

NO bc u dont want every cell replicating

base excision repair

nucleotide excision repair

NHEJ

MMR

homology directed repair

occurs during s and g2 phases only

recombination

creates variation during M1

• connections between homologous pairs provides tension necessary for meiosis

• may occur within a gene

homologous recombination

eukaryotes vs prokaryotes

topo 1 vs topo 2

ssDNA vs dsDNA cuts

what happens if ssBP not present

degrades/ coils ssDNA

homologous recomb vs HDR

transposition

jumping genes

transposase —> loops and ds break

transposon in donor chr has ds break repair via nhej

new loc is cut and transposon is added

piRNAs

silence transposable elements (help maintain genomic stability)

longer than miRNAs

transcribed from uni/ bi directional transposons

bound by piwi proteins

1. allow transposon RNA degradation (post-transcriptional silencing)

2. can direct dna methylation of transposon + repetitive elements

• important in germ line + stem cells

• role in fertility and cancer

innate immunity

adaptive immunity

antibody composure

RSS (rexombination signal sequence)

RAG complex

mechanism of recombination