BCFP S01 + S02

S01: DNA Structure and Replication; S02: Chromatin

List the pyrimidines vs purines

What is the main difference between the 2?

Pyrimidines: cytosine, thymine

Purine: adenine, guanine

Pyrimidines have 1 ring, purines have 2 rings

Identify the ribose vs deoxyribose:

Which group on a nucleotide is key for polymerization?

The 3’ OH group

The following are names of _______:

adenosine

guanosine

cytidine

uridine

thymidine

nucleosides

Nucleotide vs nucleoside

Nucleotide: base + sugar + phosphate

Nucleoside: base + sugar

Which bond links the backbone of nucleotides together? What type of bond is it?

Phosphodiester bond - covalent bond

C-G bonds contain ___ H bonds and are (more/less) stable.

A-T bonds contain ___ H bonds and are (more/less) stable.

C-G: 3 bonds; more stable

A-T: 2 bonds; less stable

Which bonds links bases together (A-T and C-G)?

Hydrogen bonds

Function of the major groove

Where transcription factors interact with DNA

In phosphodiester bonds, the ___ -OH of one nucleotide attacks the ____ -PO4 group of the next.

3’ OH group

5’ PO4 group

Structural difference between thymine and uracil

Thymine has a methyl group on its 5’ C

RNA is typically found as (shorter/longer), (ss/ds) strands.

shorter

ss

T/F: DNA replication is bidirectional with 1 origin of replication

F: bidirectional with multiple origins of replication

DNA pol II reads in the ______ direction and synthesizes in the ____ direction.

Reads 3’ to 5’

Synthesizes 5’ to 3’

Explain how DNA replication occurs in the 5’ to 3’ direction on both strands using Okazaki fragments.

DNA primase:

1. Function

2. Directionality

DNA primase lays down an RNA primer in the 5’ to 3’ direction to the newly synthesized strand, since DNA pol needs an free 3’ OH group to build off from

DNA polymerase function

Synthesizes complementary strand in the 5’ to 3’ direction

DNA polymerase 3’ to 5’ exonuclease activity

Proofreads and fixes strands in the opposite direction of synthesis

DNA helicase

Motor proteins that hydrolyze ATP to unwind DNA

Replication fork

Which enzyme creates it?

Y shaped region in DNA where it is being unwound

DNA helicase

ss DNA binding proteins

Stabilize the unwound DNA to prevent re-annealing

DNA ligase

Seals “nicks” in DNA after DNA polymerase fills the gaps left by RNA primers

Supercoiling

+ vs - supercoiling

Extra twists on the DNA strand

+ supercoiling: when DNA is overwound, making DNA strands harder to separate

- supercoiling: when DNA is underwound, allowing the helix to open up

In DNA with free ends, we need to unwind ___ DNA base pairs (aka __ helical turn) to rotate and alleviate the stress.

In DNA with fixed ends (like in our bodies), we need to unwind ___ DNA base pairs to form one ______.

In DNA with free ends, we need to unwind 10 DNA base pairs (aka 1 helical turn) to rotate and alleviate the stress.

In DNA with fixed ends (like in our bodies), we need to unwind 10 DNA base pairs to form one supercoil.

Topoisomerase

Relieves torsional stress (aka supercoiling) in DNA by cleaving bonds

Topoisomerase I vs II

I: breaks one strand and then seals

II: breaks both strands and then seals

______ DNA replication enzymes are clinical targets for anti-bacterials by (enhancing/inhibiting) strand cutting.

topoisomerase

inhibiting

______ DNA replication enzymes are clinical targets for anti-cancer by (enhancing/inhibiting) strand cutting.

Topoisomerase

enhancing

Telomerase is a (DNA/RNA) dependent (DNA/RNA) polymerase that extends _______ repeats to the _____ strand.

Telomerase is an RNA-dependent DNA polymerase that extends telomere repeats to the parental strand.

Telomere

Protective repetitive stretches of DNA at the tips

What type of enzyme is telomerase? Explain how it functions.

Reverse transcriptase: contains RNA templates and synthesizes DNA complementary to its own RNA. This gets added to the 3’ of the parental strand so the ends of DNA can be replicated.

End-replication problem

On the end of lagging strand, there is no free 3’ OH group for DNA polymerase to build off of. This would cause the chromosome to shorten a little bit during every replication.

Chromatin = ____ + _____ proteins + _____ proteins

DNA + histone + non histone proteins

List if the following events require mostly euchromatin or heterochromatin:

Transcription

Mitosis

G1/S/G2

Transcription: euchromatin

Mitosis: heterochromatin

G1/S/G2: euchromatin

Nucleosome structure

2 copies of each of the 4 core histone proteins with DNA wrapped around it. Beads are separated via linker DNA.

What are the 4 core histones found in nucleosomes?

H2A, H2B, H3, H4

H1 linker histone

Promotes more compact chromatin folding (10 nm → 30 nm fiber)

What are the 2 parts of a histone protein?

The histone fold and the N-terminal tail

The histone fold consists of ___ alpha helices and ___ loops that dimerize with another histone protein. These combine to form a ___mer that forms the bead of the nucleosome.

3 alpha helices and 2 loops

octomer

The N-terminal tail of core histones contain (acidic/basic) proteins that contain AAs like ____ and _____.

basic

lysine, ariginine

The ______ of core histones are the targets of post-translational modifications

N-terminal tails

_____ catalyze acetylation of lysines on core histones.

_____ catalyze deacetylations of lysines on core histones.

HATs (histone acetyltransferases) catalyze acetylation of lysines on core histones.

HDACs (histone deacetylases) catalyze deacetylations of lysines on core histones.

Generally, (acetylation/deacetylation) and (methylation/demethylation) are associated with euchromatin and increased transcriptional activity.

acetylation and demethylation

Acetylation of core histones works by ____ positively charged lysines, which ___ histone interactions with negatively charged DNA and loosens the chromatin.

neutralizing

weakens

_____ catalyze methylation of lysines on core histones.

_____ catalyze demethylations of lysines on core histones.

Histone methyltransferases (HMTs) catalyze methylation of lysines on core histones.

Histone demethylases (HDMs) catalyze demethylations of lysines of core histones.

Lysine acetylation vs methylation net charge changes

Acetylation: lysine + → neutral

Methylation: no net charge change

The histone code explains how _____ proteins are recruited to chromatin when they recognize ________ modifications such as ___ and ____.

The histone code explains how reader proteins are recruited to chromatin when they recognize post-translational modifications such as acetylation and methylation.

______ proteins are responsible for altering local chromatin structure after recruitment.

Reader

T/F: reader proteins recruitment leads to heterochromatin formation

F: depends, can lead to heterochromatin or euchromatin depending on the histone post-translational modification

(Bromodomains/chromo (PHD) domains) recognize acetylated histones.

(Bromodomains/chromo (PHD) domains) recognize methylated histones.

These are both examples of ______.

Bromodomains recognize acetylated histones.

(Chromo (PHD)) domains recognize methylated histones.

reader proteins

Constitutive heterochromatin are found in _______.

centromeric DNA

What are the 3 ways to alter chromatin structure?

1. Histone post-translational modifications

2. Chromatin remodeling complexes

3. Histone variants

Chromatin remodeling complexes

ATPases that disrupt histone-DNA contacts within nucleosomes

2 ways that chromatin remodeling complexes alter histone-DNA contacts

1. Nucleosome sliding: sliding nucleosomes to expose underlying DNA

2. Altering DNA-histone interactions (ex: to loosen and expose DNA)

Histone variants

Variants of histones that have specialized effects

Telomeres are (ss/ds) DNA that form a __-loop to _________.

Telomeres are ss DNA that form a t-loop to protect the chromosomes from being mistaken for a ds break that needs to be repaired.

Centromere

Region of chromosomal DNA where sister chromatids connect and kinetochores assemble.

Kinetochore

Protein complex that binds centromeres and acts as an attachment site for microtubules and chromosomal separation in mitosis/meiosis.

Giemsa staining (G-banding) is a (high/low) resolution method of visualizing chromosomes. It can detect (large/small) scale chromosomal features like the ____ of chromosomes.

low resolution

large scale

number

Giemsa staining (G-banding) source material AND staining method

WBCs arrested in metaphase

Stains regions with a higher AT base composition

Metacentric

Submetacentric

Acrocentric

Metacentric: centromere in middle

Submetacentric: centromere slightly closer to one end

Acrocentric: centromere significantly closer to one end

Primary constriction of chromosomes

The centromere, where the sister chromatids are most tightly connected

T/F: sister chromatids are connected along the whole length of a chromosome

T

p vs q arm of chromosome

p: short arm

q: long arm

The human genome is made up of roughly ___% single copy genes and ___% repetitive DNA sequences.

5% single copy genes

95% repetitive DNA sequences

Within the 95% repetitive DNA of a human genome, it can be divided into which 2 categories?

1. Tandem: arrays of repeated DNA sequences

2. Dispersed: DNA repeats occuring throughout the genome

Minisatellites vs microsatellite array and repeat lengths

Minisatellite: arrays of 0.1-20 kb, with repeat units of 10-70 bp

Microsatellite: arrays <100 bp long, with repeat units of < 10 bp

Minisatellites are also called ______.

Microsatellites are also called _____.

Mini: VNTRs

Micro: STRPs

Expansion of trinucleotide repeats, present in Huntington’s, myotonic dystrophy, and Fragile X syndrome

Trinucleotide repeat syndromes

Mini/microsatellites are examples of (tandem/dispersed) repeats.

tandem

LINES and SINES are examples of (tandem/dispersed) repeats.

dispersed

Explain what it means that LINES/SINES are retrotransposons.

They can copy and paste themselves elsewhere into DNA.

Explain how LINES are autonomous.

1. LINE is transcribed into RNA

2. This RNA encodes a reverse transcriptase that allows them to synthesize their own DNA and insert it elsewhere.

SINES are (autonomous/non-autonomous), which means ______.

non-autonomous

they use LINE elements to form a reverse transcriptase

SINES are ______ bp in length.

LINES are ______ bp in length.

SINES: 80-400 bps

LINES: several kilobases

Much of the human genome variation occurs in the sequence and repeat numbers of _____.

VNTRs (minisatellites) and STRPs (microsatellites)