DNA & RNA Organization

What enzymes break glycosidic linkages?
glycolases
What enzymes cleave the phosopho-ester linkage?
nucleases

How many H bonds are between A:T/U?

G:C?

2,3
A double helix is stabilized by what forces?
H bonds between bases; hydrophobic forces; van der Waals interactions
What bonds are broken during denaturation of DNA?
Only H bonds between strands.
At what wavelength do nucleobases absorb light? What would this absorption show with respect to DNA structure?
260 nm; denaturation
What is DNA melting or transition temp? What increases melting temp?
Temp at which 50% of the helix is denatured; Melting temp increases with GC base content
tRNA are about how long and make up what percent of total cellular RNA?
75-85 bp; 10-15%
Which RNA contain several unusual bases such as pseudouridine and dihydrouridine?
All tRNA.
What role do tRNAs play?
They serve as adapter molecules, carrying specific AAs to the site of protein synth. There is at least 1 tRNA/AA.
Which RNA has the highest percentage of modified bases?
tRNA
The 3′ end of what RNAs have the sequence CCA? Where does the AA attach to this RNA? What this AA attached RNA called?
All tRNAs; 2’C or 3’C of the sugar of the last nucleotide; aminoacyl-tRNA
Which RNAs are most abundant in the cell?
rRNA
How are the sizes of rRNAs measured? What are the measurements in pro and eukaryotes?
Based on sedimentation coefficient; Pro: 23S, 16S, and 5S; Euk: 28S, 18S, 5.8S, and 5S
What fraction of a ribosome is made up of rRNA?
2/3 rRNA; 1/3 protein
What do 23S and 28S rRNA do? What does 16S do?
Catalyze the peptide bond formation (ribozyme); identifies the first codon during translation
What percent of the cellular RNA is mRNA? Describe the size and structure of mRNA.
2-5%; Size is highly variable. Little intrastrand bp. Eukaryotic mRNA is capped at 5′ end (with 7-methylguanosine triphosphate attached to C5′ of first nucleotide via an unusual 5-5 phosophodiester link) and has poly(A) tail at 3′ end. Prokaryotic is monocystronic while eukaryotic is poly cystronic.
hnRNA (heterogeneous nuclear RNA)
precursors of mRNA with 5′ cap and poly(A) tail
snRNA (small nuclear RNA)
90-several hundred nucleotides long; rich in U; involved in maturation of pre-mRNA
scRNA (small cytoplasmic RNA)
associated with proteins involved with recognition of signal peptides on proteins
**miRNA (micro RNA)**
19-25 nucleotides long; noncoding regulatory RNA; ~750 identified in humans; function in post-transcriptional gene silencing by inducing target RNA degradation or translational inhibition; crucial regulators of gene expression; may find application as cancer biomarkers, in gene therapy, and treating infx in the future.
DNA double helix is found primarily in what form? Briefly describe the other 2 forms.
B form; A form has right turn and DNA:RNA hybrid. Z form has left turn and alternating purine and pyrimidine stretch. All forms can be found in the same molecule of DNA.
Describe prokaryotic DNA.
Plectonemic; negatively supercoiled (by gyrase); covalently closed circular DNA;;
What 2 structures result from supercoiling?
plectonemic (intertwined) or solenoidal (spring-like)
What is gyrase?
A topoisomerase – the only one capable of introducing negative coiling; ATP dependent; makes transient double stranded breaks; the target of anti-bacterial drugs like cipro and novobiocin.
Describe eukaryotic DNA structure.
Chromosomal DNA is linear and mitochondrial is circular. Not double stranded all the way to the end as the 3′ end is longer (3′ overhang). Both ends have long segments of repeated hexameric sequence called telomers (~250-1500 hexameric repeats which varies from species to species).
Nonrepetitive sequence of DNA
Only one copy per haploid genome and codes for proteins.
Describe the 3 types of DNA repetitive sequences.

1. Highly repetitive – 5-500 bp repeated many times in tandem, found in centromeres ; telomeres; forms minisatelites.

2. Moderately repetitive – ;10^6 copies/genome, dispersed, usually made of transponsons (long and short interspersed rpt sequences (LINEs and SINEs)

3. Microsatellite repeat sequences – 2-6 bp repeats, rpt 2-50 times in tandem at several locations in the genome, responsible for microsatellite polymorphism, trinucleotide rpts are involved in disease causing mutations.

Histones
Structural proteins around which eukaryotic DNA is negatively supercoiled, forming solenoidal super coils. Rich in Lys and Arg; histone complex is called chromatin; 5 histones – H1, H2A, H2B, H3 and H4; DNA wraps histone in ~2 turns (~150 bp); Histone core-DNA complex is a nucleosome; spacer region is 30-50 bp complexed with H1
Name the 2 functional states of chromatin. How is chromatin structure altered (2 ways)?

Euchromatin (lightly condensed) and heterochromatin (highly condensed);

1. Methylation of C residues leads to chromatin compaction and gene silencing.

2. Histone modification (e.g. phosphorylation or acetylation – most common)

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