Detailed explanation of rna metabolism lecture note

1. MEDICAL BIOCHEMISTRY,
FACULTY OF BASIC MEDICAL
SCIENCES
RNA METABOLISM I
ISHAKA AMINU
MBBS, PHD.

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Outline
1. Introduction
2. Structure of RNA
3. Types of RNA
4. Transcription: requirements, structure of
RNAP II, steps.

3. ▪ Ribonucleic acid (RNA), macromolecule for storage and transmission
of genetic information
▪ It provides link between gene encoded in DNA and the actual
cellular activities
▪ On average gene is formed by 10-50,000 bp, longest is 2.5m bp.
▪ Found in nucleus, organelles such as ribosome, and cell vacuoles
fluid
▪ Ribozyme discovery, extends the definition of enzymes beyond
being only proteins, RNA also has catalytic activity
▪ Thus, RNA has both structural and catalytic roles.
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Introduction

4. Central dogma of
molecular biology
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Introduction

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▪ Single stranded structure
▪ Contains ribose (DNA 2’-deoxy ribose)
▪ Contains uracil base (DNA Thymine)
▪ Does not obey Chargaff’s rule
▪ Susceptible to alkaline hydrolysis
▪ Retains all genetic information of DNA
▪ Specific RNA for specific functions; thus,
Characteristically diverse (many types)
▪ Structure similar to A-form DNA
Structure of RNA

6. ▪ There are 3 major types
1. Transfer RNA (tRNA): 10-20%, carries amino acids to translation
machinery, it is very stable
2. Ribosomal RNA (rRNA): 50-80%, makes up majority of cellular RNA,
much in the ribosomes, also very stable.
3. Messenger RNA (mRNA): 5-10%, Encodes messages from DNA to
ribosomes, unstable as is rapidly degraded by nucleases
▪ Other RNAs
1. Heterogeneous nuclear RNA (hnRNA): a precursor for mRNA and
other RNAs
2. Small nuclear RNA (snRNA): involved in mRNA processing
3. Small nucleolar RNA (snoRNA): role in rRNA molecules processing
4. Small cytoplasmic RNA (scRNA): involved in selection of proteins for
export
5. Transfer-messenger RNA: present in bacteria, adds short peptide
tags to proteins to facilitate degradation of abnormally synthesized
proteins
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Types of RNA

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▪ RNA synthesis is similar to DNA replication in
▪ Directionality (5’→3’), however, is DNA-directed
▪ Steps: initiation, elongation, and termination
▪ Copied from a template
▪ Large multicomplex initiation
▪ Different from DNA replication in
▪ No primer is required
▪ U in place T base, ribose
▪ Only discrete segments of DNA template copied (very
small portion= gene)
▪ Only one strand of DNA serves as template
▪ Different set of enzymes required
▪ No proofreading
▪ Doesn’t stop at one cycle
RNA synthesis: Transcription

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Part of transcribed RNA that results in protein
▪ Close to 96% of RNA in higher
eukaryote is not translated into
protein
▪ Out of these, 50-70% are introns
(non-coding sections of RNA)
▪ Only about 4% form the coding
segment

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Requirements in transcription
A. Template
B. Substrates: ATP, GTP, CTP, UTP, and Mg2+
C. Enzyme: DNA dependent RNA polymerase
▪ RNA polymerase (RNAP) is of 3 types
1. RNAP type I (A): responsible for rRNA synthesis. Not
sensitive amanitin (a toxin in mushroom).
2. RNAP type II (B): main enzyme that catalyzes the
synthesis of mRNA.
▪ It has 5 subunits
▪ Activated by phosphorylation
▪ Inhibited by alpha amanitin
▪ It blocks translocation of the enzyme
3. RNAP type III (C): responsible for the synthesis of tRNA.
Moderately sensitive amanitin

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Structure of RNAP II (B)
▪ Complex cos has to do all jobs on
its own
▪ 5 sub-units
▪ 2 α : CTD, NTD- αCTD
recognizes the UP element on
the promoter; αNTD holds
αCTD via a thin loop, and β
sub-unit
▪ β and β’ carry the catalytic
activity domain σ
▪ σ is the most important sub-
unit which attaches to the
promoter sequence region
▪ Promoter region is TATA box and
CAAT box
β β'

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Transcription-initiation stage
▪ σ Sub-unit of RNAP II attaches to
promoter region on
DNA:TATA/Prinbow (Prokaryotic -10
to -35 bp)/Golberg-Hogness
(Eukaryotic-25 to -30 bp) box and
CAAT box
▪ Transcription factors first recognize
and bind to promoter region at the
start point
▪ Humans have 100,000 transcription
initiation sites
▪ RNAP separate the 2 strands apart
within the gene
▪ The unwinding of DNA is also assisted
by topo-isomerase.
Promoter
region
RNAP
Non-template
(sense) strand
Template (Antisense)
strand

12. ▪ Elongation goes until the entire
gene is transcribed
▪ In the process, ribose(not
deoxyribose) and Uracil (not
thymine) containing nucleotides
are synthesized
▪ 5’→ 3’ mRNA is produced
▪ RNAP II zips back DNA as it
goes, keeping only 10-20 bases
exposed at a time
▪ The region of unpaired DNA due
to the action of RNAP II is called
transcription bubble
▪ Due to absence of proofreading,
there are 10,000-100,000
chances of mistakes in
transcription than in DNA
replication
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Transcription-elongation stage

13. ▪ Once RNAP reaches the end of
the gene, termination occurs
due to signal by a specific
protein called ‘rho factor’ (ρ) an
ATP dependent.
▪ RNAP detaches itself since it
cannot go further due to rho
factor
▪ mRNA produced leaves
▪ DNA returned to it original
structure
▪ Note: rho independent
termination has also been
described.
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Transcription-termination stage

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Post-transcriptional processing as
RNA metabolism II

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