In the chromatin structure, which histones form the octamer, and which ties the nucleosome beads together in a string?
octamer = 2 sets of H2A, H2B, H3, H4; H1 ties nucleosome beads together
Octamer subunits consist primarily of which 2 amino acids?
lysine and arginine
Which type of chromatin is condensed, transcriptionally inactive, and sterically inaccessible?
HeteroChromatin = HighlyCondensed
Which type of chromatin is less condensed, transcriptionally active, & sterically accessible?
Euchromatin; Eu=tru; "truly transcribed"
What does PURe As Gold stand for?
Purine nucleobases = Adenine & Guanine
What does "CUT The PY" stand for?
PYrimidine nucleobases = Cytosine, Thymine, & Uracil
How many rings do purine hav?
How many rings do pyrimidines have?
How does cytosine become uracil?
Deamination of cytosine makes uracil
What polar group does guanine have?
What non polar group does thymine have?
Which bond is stronger, AT or CG? Why? What is the laboratory consequence of this?
CG (3 H-bonds) > AT (2 H-bonds); An ↑ in CG bonds will ↑ melting temperature
What 3 amino acids are necessary for purine synthesis?
Glycine, aspartate, & glutamine
What makes of a nucleoside?
Base + ribose = Nucleoside
What makes up a nucleotide?
base + ribose + phosphate = Nucleotide (linked by 3'-5' phosphodiester bond)
What are purines made from?
What are pyrimidines made from?
Orotate precursor, with PRPP (phosphoribosyl pyrophosphate) added later
What is made first, ribonucleotides or deoxyribonucleotides? What enzyme is necessary to convert one to the next?
ribonucleotides are synthesized first and are converted to deoxyribonucleotides by ribonucleotide reductase
What 2 pathways is carbamoyl phosphate involved in? What enzyme deficiency leads to a build up of carbamoyl phosphate which is then converted to:__?
Carbamoyl phosphate is involved in: (1) de novo pyrimidine synthesis & (2) the urea cycle. Ornithine transcarbamoylase deficiency (urea cycle) leads to build up of carbamoyl phophate which is then converted to orotic acid.
Name 5 drugs that interfere with nucleotide synthesis:
(1) hydroxyurea: inhibits ribonucleotide reductase (2) 6-mercaptopurine: blocks de novo purine synthesis (3) 5-fluorouracil: inhibits thymidilate synthase (4) methotrexate: inhibits DHFR (5) Trimethroprin: inhibits bacterial DHFR
Inhibition of thymidilate synthase & inhibition of dihydrofolate reducatse (DHFR) leads to a decrease in what substance?
Which step in the de novo purine & pyrimidine synthesis pathway requires just aspartate?
orotic acid to UMP
Which step in the denovo purine & pyrimidine synthesis pathway requires aspartate, glycine, glutamine, & THF?
Ribose 5-P to PRPP
An inability to convert orotic acid to UMP would be caused by a deficiency of either of which two enzymes? What is this disease called and what is its inheritance?
orotic acid phosphoribosyltransferase or orotidine 5-phosphate decarbosylase; Dx = orotic aciduria (autosomal recessive)
What are the clinical findings in orotic aciduria?
↑ orotic acid in urine; megaloblastic anemia (does not improve w/ administration of B12 or folic acid); failure to thrive; NO hyperammonemia (vs OTC deficiency)
What lab value can distinguish orotic aciduria from ornithine transcarbamoylase def?
OTC has hyperammonemia; orotic aciduria does not
What is the tx for orotic aciduria?
oral uridine administration
What does adenosine deaminase deficiency cause a build up of? What does this lead to?
excess ATP & dATP imbalances nucleotide pool via feedback inhib of ribonucleotide reductase; this prevents DNA synth & ↓ lymphocyte count (a major cause of SCID)
Adenosine deaminase def is an important cause of what immunodeficiency?
↓DNA & ↓lymphocytes → SCID
Lesh-Nyhan is a defect in which enzyme? What are the biochemical results?
Defect in purine salvage d/t NO HGPRT; excess uric acid production
What is the enzyme HGPRT responsible for?
HGPRT converts: hypoxanthine → IMP & guanine → GMP
How is Lesh-Nyhan inherited? What are the clinical findings?
X-linked recessive; retardation, self-mutilation, aggression, hyperuricemia, gout, choreoathetosis
What reaction does adenosine deaminase normally catalyze?
Adenosine → Inosine
What does an unambiguous genetic code refer to?
each codon specifies only 1 amino acid
What does degenerate/redundant genetic code refer to? What is the exception to genetic redundancy?
more than 1 codon may code for the same amino acid; Methionine is the exception: it is encoded by only 1 codon (AUG)
What does commaless, nonoverlapping genetic code refer to?
read from a fixed starting point as a continuous sequence of bases
What does universal genetic code refer to and what are some exceptions?
genetic code is conserved throughout evolution, except for mitochondria, archaebacteria, mycoplasma, and some yeast
What is a silent mutation?
same AA, often base change in 3rd position of codon (tRNA wobble)
What is a missense mutation?
changed AA (convservative - new AA is similar in chemical structure)
What is a nonsense mutation?
change resulting in early stop codon
What is a frame shift?
change resulting in misreading of all nucleotides downstream, usually resulting in a truncated nonfxnal protein
What is the order of severity for the different types of mutations?
nonsense > missense > silent
Eukaryotic and prokaryotic DNA synthesis is _1_ and involves both _2_ and _3_ synthesis.
1: semiconservative; 2: continuous; 3: discontinuous (Okazaki fragment)
For eukaryotes, where does replication begin?
begins at a consensus sequence of base pairs
What is the origin of replication?
A particular sequence of DNA where replication begins - may be single or multiple
Describe the replication fork
Y shaped region along the DNA template where leading and lagging strands are synthesized
What does helicase do?
unwinds DNA template at replication fork
What do the single stranded binding proteins do?
SS binding proteins prevent strands from reannealing
What do DNA topoisomerases do?
DNA topoisomerases create a nick in the helix to relieve supercoils created during replication
What does primase do?
Primase makes RNA primer on which DNA poly III can initiate replication
What does DNA polymerase III do?
prokaryotic only - elongates leading strand by adding deoxnucleoTIDEs to the 3' end. Elongates lagging strand until it reaches primer of preceding fragment. 3' → 5' exonuclease activity "proofreads" each added nucleoTIDE
DNA Polymerase III has 5'→3' (__1__) and proofreads with 3'→5' (__2__).
1: synthesis; 2: exonuclease
What does DNA polymerase I do?
prokaryotic only - degrades RNA primer and fills in the gap with DNA
DNA Polymerase I (__1__) RNA primer with 5'→3' (__2__).
1: excises; 2: exonuclease
What does DNA ligase do?
How do fluoroquinolones work? (Abx)
inhibit DNA gyrase specific for prokaryotic topoisomerase
In which single strand DNA repair mechanism do specific endonucleases release the oligonucleotide containing damaged bases and DNA polymerase and ligase fill and reseal the gap?
Nucleotide Excision Repair
What disease occurs when there is a genetic defect in nucleotide excision repair? Describe the disease and its inheritance
Xeroderma Pigmentosum, prevents repair of thymidine dimers caused by UV light; Autosomal Recessive
In base excision repair, (__1__) recognizes and removes damaged bases and (__2__) cuts the DNA at (__3__) site to remove the empty sugar; then the gap is rilled an resealed.
1: specific glycosylases; 2: AP endonuclease; 3: apyrimidinic site
Which SS DNA repair mechanism does this describe? - Unmehtylated, newly synthesized string is recognized, mismatched nucleotides are removed, and the gap is filled and resealed.
Mismatch repair is defective in which inherited disease?
Hereditary nonpolyposis colorectal cancer (HNPCC)
Is there any requirement for homology in nonhomologous end joining of ds DNA repair?
no: it's non homologous; brings together 2 ends of DNA fragments
In what direction are DNA and RNA both synthesized?
Which carbon bears the triphosphate and the energy source for bond formation?
The 5' end of the incoming nucleotide
What is the target of the 3' hydroxyl attack?
The triphosphate bond
In which direction is protein synthesized?
N to C
What is the most abundant type of RNA?
What is the longest type of RNA and what is the shortest type of RNA?
What does the start codon (AUG) code for in eukaryotes & prokaryotes?
euk: methionine; prok: formyl-methionine (f-Met)
What are the mRNA stop codons?
UGA, UAA, UAG
Which part of the DNA binds RNA polymerase and multiple other TFs upstream from gene locus?
Promotor- TATA box & CAAT box; AT rich sequence
What is the stretch of DNA that alters gene expression by binding of transcription factors?
This is the site of DNA where negative regulators bind:
A promoter mutation commonly results in a dramatic (__) in the amount of gene transcribed.
A dramatic decrease in the amount of gene transcribed
In eukaryotes, what does RNA polymerase I make
In eukaryotes, what does RNA polymerase II make?
In eukaryotes, what does RNA polymerase III make?
Which RNA polymerase opens DNA at promotor site?
RNA poly II
RNA polymerase can't proofread, but what can it do?
initiate chains; RNA polymerase II opens DNA at promoter site
In prokaryotes, what makes the different types of RNA?
1 kind of RNA polymerase with multiple subunit complex makes all 3 kinds of RNA
Why does alpha amanitin cause liver failure and where is it found?
inhibits RNA polymerase II, found in death cap mushrooms
What 3 steps in RNA processing occur after transcription?
(1) capping on 5' end (7-methyguanosine) (2) polyadenylation on 3' end; ~200 A's (3) splicing out of introns
What is the initial transcript called and what is the capped and tailed transcript called?
Initial transcript: heterogeneous nuclear RNA (nhRNA); capped & tailed transcript: mRNA
Which enzyme involved in RNA synthesis does not require a template? What is the signal?
poly A polymerase; polyadenylation signal is AAUAA
What kind of RNA is transported out of the nucleus?
only processed RNA
Describe the 3 steps in pre-mRNA splicing of eukaryotes.
(1) Primary transcript combines w/ snRNPs & other proteins to form spliceosome. (2) Lariat shaped intermediate is generated. (3) lariate is released to remove intron precisely & join 2 exons
Patients w/ lupus make antibodies to ____.
Make antibodies to spliceosomal snRNPs
What part of the pre mRNA contains the actual genetic information coding for protein?
How many nucleotides is a tRNA & what form does the secondary structure take?
75-90 nucleotides; Cloverleaf form
Which end of the tRNA is the amino acid bound to?
covalently bound to the 3' end (w/ CCA)
What does aminoacyl tRNA synthetase do to the AA before and after it binds to tRNA, and what happens if it's the wrong match?
Scrutinizes. Hydrolyzes the bond if it is the wrong match
What does a mischarged tRNA do?
reads usual codon but inserts wrong amino acid
How to tetracyclines work?
bind 30s subunit, preventing attachment of aminoacyl-tRNA
Accurate base pairing is required only in the first two nucleotide positions of an mRNA codon, so codons differing in the 3rd position may code for the same tRNA/AA due to degenerate genetic code. What is this called?
What initiates protein synthesis?
GTP hydrolysis, initiation factors (eIFs) help assemble 40S ribosomal subunit with initiator tRNA and are released with mRNA and the ribosomal subunit assemble with the complex
What happens in elongation of protein synthesis?
Aminoacyl-tRNA binds A site (except for initiator methionine); ribosomal rRNA catalyzes peptide bond formation, transfers growing peptide to AA in A site; ribosome advances 3 nucTIDE toward 3' end of RNA, moving peptidyl RNA to P site (translocation)
What happens in termination of proteins synthesis?
stop codon is recognized by release factor, and completed protein is released from ribosome
What ribosomes do eukaryotes have?
40S + 60S → 80S (Eukaryotes, Even)
What ribosomes do prokaryotes have?
30S + 50S → 70S (prOkaryotes, Odd)
What is the energy source for tRNA activation?
What is the energy source for translocation?
GTP (gripping & going places)
How do aminoglycosides work?
by inhibiting formation of the initiation complex and cause misreading of mRNA
How does chloramphenicol work?
inhibits 50S peptidyltransferase?
How do macrolides and clindamycin work?
binds 50S, blocking translocation
What is trimming in posttranslational modifications?
removal of N or C termal propeptides from zymogens to generate mature proteins
What are covalent alterations in posttranslational modifications?
phosphorylation, glycosylation, hydroxylation
What is proteasomal degradation in posttranslational modification?
attachment of ubiquitin to defective proteins tag them for breakdown