IAS01 Flashcards
pentose of nucleotide characteristics
1’: link to base
2’: determine DNA or RNA
3’: link to phosphate of adj nucleotide
5’: link to phosphate
DNA v RNA
DNA: DS, 2’ hydrogen, thymidine, for info storage, resilient (permanent)
RNA: mostly single stranded, 2’ -OH, uridine, various functions, transient
nucleotide bases & base pairing
adenosine – uridine / thymidine
cytidine — guanosine
purine - pyrimidine
note: T +1 -CH3 group from U
chargaff’s rule: %A=%T, %C=%G
DNA structure & grooves
rh double helix, ds, read from 5’ to 3’, antiparallel, 10.4 base per turn
base in center, backbone at outside
major grooves & minor grooves, major more accessible to transcription factor binding as less distortion of DNA shape
RNA structure
mostly single-stranded, more struc variety, more varied functions, rarely base pairing
can form more H bonds by binding to RNA nts apart from base pairing
RNA functions & types
info transfer (mRNA)
AA carrier (tRNA)
catalyst (rRNA)
sgRNA, snRNA, siRNA
DNA coiling
coil around +vely charged histone octamers, namely H2A, H2B, H3, H4, to form nucleosome (basic subunit of chromatin) -> fiber -> loop (75k nt) -> rosette (6 loops) -> coil (30 rosettes) -> chromatid (10 coils)
central dogma of molecular biology & newer developments
DNA -> RNA -> protein i.e. replication, transcription, splicing, translation
HIV virus do reverse transcription
but cannot return from protein to mRNA
collinearity
relationship between DNA base sequence & protein AA sequence
Sense/coding strand (5’ → 3’) = mRNA stand (5’ → 3’) = polypeptide (N->C)
transcription initiation
TATA-box binding protein binds to TATA box of promoter
other components of TFII bind i.e. transcription factors assemble at promoter
mediator carries RNA polymerase II to promoter –>
combine to form transcription initiation complex
transcription elongation
helicase unwinds DNA to expose base in transcription bubble
antisense / template strand act as template for RNA synth which is read in 3’ to 5’, sense / coding strand not involved & go outside
free RNA nt triphosphate enters RNAP -> hydrolyze to form RNA nt & bind to template strand one at a time -> hybrid helix forms -> nascent / pre-mRNA forms 5’ to 3’ & exits in diff. strand
transcription termination, requirement & energy source
terminator sequence or randomly
Mg2+ dependent
energy from ATP & hydrolysis of RNA TPs to move RNAP & form mRNA chain
splicing major processes
5’ capped (5’-5’ bond) to stabilize DNA
3’ end polyadenylated
introns removed & cleaved by spliceosome (protein complex) & exons remain
edited mRNA travels out of nucleus by pore to cytoplasm
splicing (OPTIONAL)
spliceosome removes 1 intron
Assembly proteins assemble at intron/exon borders, U1 binds to 5’ end, U2AF & BBP binds to 3’ end, splicing factors act as beacons to guide 5 snRNP, U2, U4, U5, U6, to promote spliceosome formation
Spliceosome brings exons on both intron ends close together
Intron end cuts 5’ end at GU & folded back on itself to A -> loop / intron lariat
Spliceosome cuts 3’ end at AG -> detach intron -> exons joined / splice sites connected
mRNA released, spliceosome disassembles
alternative splicing
diff. splicing patterns
exons joined diff. comb. OR diff. exons used in diff. proteins
-> diff. proteins from 1 pre-mRNA
SMA genetics
originated from issues in splicing (autosomal recessive)
SMN2 C->T prevents binding of helper protein in intron -> spliceosome cannot assemble -> exon 7 removed along w/ introns -> protein shorter than normal i.e. alternative splicing (7/8 exons remain)
SMN nonfunctional -> not ensure survival of motor neuron -> motor neuron die
SMA symptoms
muscle wasting, weakness of muscle, infant death
genetic code properties
triplet code (1 codon, 3 nt, 1 AA)
degenerate: multiple codons code for same AA
existence of stop codons w/o tRNA
ribosome structure
rRNA held in place by proteins
large & small subunits, 5’ E P A 3’ site
large: catalyze peptide bond formation
small: mRNA positioning to read as codons i.e. mRNA reading
tRNA
adaptor btn mRNA & protein
carries individual AA, 20 types each carrying 1 type of AA
(charging: binding of tRNA w/ AA by aminoacyl tRNA synthetase)
translation initiation
small sunbunit binds to mRNA & moves to 1st codon -> tRNA carrying Met binds to AUG codon -> large subunit binds to small subunit to form complex while tRNA at P site
translation elongation
tRNA binds A -> P -> E (exit) site
A: match anticodon w/ codon, growing chain in P site added to AA in A site by peptide bond catalyzed by ribosome -> moives to P by ribosome moving 3nt across
P: AA removed i.e. deacylated -> moves to E
E: ejected & recycled
translation termination
stop codon reached, no tRNA & instead recruit release factor -> AA chain & mRNA released -> ribosome disassemble
histone posttranslational modification
methylation, acetylation, phosphorylation, etc. change histone struc, affects part of DNA exposed in nucleosome -> affect gene expression
transcription factors
bind to DNA promoters to trigger transcription
DNA enhancer regions & insulator regions
enhancers increase chance of transcription:
cis-acting enhancers few knt away bind to activator, forms loop -> attach to TIC to trigger transcription
insulators reduce chance of transcription:
inhibitor protein e.g. CTCF bind to insulator to prevent TIC formation
DNA methylation & e.g. in gene silencing & activation
e.g. silences gene as transcription factors cannot bind to methylated protein
e.g. in DNA insulator silencing, methylation of C prevent CTCF binding, cannot prevent TIC formation
mediator (transcription)
binds several transcription factors & controls transcription, can enhance or silence gene
demonstrates complexity as multiple TF needed to activate mediator
gene expression regulation
histone posttranslational modification
enhancers, insulators, transcription factors
posttranscription processing & posttranslational processing control
mature mRNA degrade into nt
modified protein degrade into AA
cell variety & gene expression
diff. proteins expressed in diff. cells & diff. dev. stages for diff. function due to gene expression
failure to do so lead to cancer
reverse transcription application
reverse transcription of mRNA into cDNA analyzes mRNA expression in cancer cells (overexpression or underexpression) by fluorescing cDNA & adding into microarray
compare amount of cDNA reversely transcripted from one sample w/ cDNA of another sample (w/ CBP)
used in diagnosis & management of cancer
