Download A. Introduction

TRANSCRIPTION (RNA SYNTHESIS)
I.
INTRODUCTION
A. Coding and Template DNA strands
1. Coding DNA
a) This is the DNA strand that is complimentary to the DNA strand being transcribed
(1) It's sequence will be identical (not complimentary) to the RNA molecule synthesized from the
coding strand
(a) Except Ts replace Us
b) The coding strand sequence is given when talking about double stranded DNA
(1) Same polarity as RNA
(2) Easier when referring to the genetic code
(3) Always given in the 5' to 3' direction
2. Template strand
a) This is the strand of DNA that RNA polymerase binds to during transcription
b) It is complimentary to both the coding strand and the RNA that is transcribed from it
3. A single strand of DNA in a chromosome will have some regions that are the coding
strand and other regions in which it is the template strand
a) Some viruses, to conserve genome size, will have an area of double-stranded DNA
in which each strand is a coding strand
B. Basic feature of RNA synthesis
1. Substrates
a) DNA-dependent RNA polymerase
b) Four ribonucleoside triphosphates
(1) ATP, GTP, CTP, and UTP
c) DNA template
(1) The enzyme that performs transcription is referred to as a DNA-dependent RNA polymerase
(a) Here we will refer to it simply as RNA polymerase
d) No primer
(1) No primers are needed for initiation of RNA synthesis which is in contrast to DNA polymerases
2. Reaction
a) 3'-OH group of one nucleotide reacts with 5'-triphosphate of another [(NMP)n + NTP
 (NMP)n+1 + PPi]
(1) Pyrophosphate is cleaved
(2) This is similar to DNA synthesis
(3) This reaction results in the formation of a phosphodiester bond
b) Catalyzed by RNA polymerase
3. Complimentary and antiparallel
a) The sequence of bases in RNA is determined by the base sequence of DNA in a
complimentary mode
(1) The ribonucleotides being added to the RNA molecule must H-bond properly to the
deoxyribonucleotides of the template DNA strand
(a) dA-U, dG-C, dT-A, dC-G
b) Antiparallel
(1) RNA is synthesized 5' to 3' from a DNA molecule going 3' to 5'
(a) RNA molecule has 5'-triphosphate and a 3'-OH ends
C. Stages of transcription
1. Initiation
a) Binding of RNA polymerase to DNA promoter sequence
2. Chain elongation
a) Polymerization of RNA
3. Termination and release
a) Removal of RNA polymerase from DNA and the release of the RNA molecule
II. INITIATION
A. Introduction
1. RNA polymerase must bind to the promoter, separate the strands of DNA, and find
complimentary ribonucleotides to the DNA bases
B. Promoter and -35 region
1. RNA polymerase binding to a region of DNA referred to as the promoter
2. Numbering
a) The first base of the first codon transcribed is given the number +1
b) Downstream is the direction of transcription
c) Upstream bases, which are not transcribed, are given negative numbers
3. Sequence of promoter
a) Pribnow box
(1) Sometimes referred to as the TATAAT box
4. There is also another conserved sequence further upstream
a) -35 sequence
III. CHAIN ELONGATION
A. Definition
1. After about 8 ribonucleotides are added to the RNA molecule, RNA polymerase
undergoes a conformation change and loses the sigma factor
a) This marks the beginning of the elongation phase of transcription
b) Elongation is catalyzed by the core enzyme
(1) The holoenzyme lacking the sigma factor
2. RNA polymerase proceeds down the DNA molecule, separating DNA strands, binding
ribonucleoside triphosphates that will H-bond with the template strand, and adding them
to the growing RNA chain
a) The RNA strand dissociates from DNA and the DNA double-helix reforms behind
RNA polymerase
B. Rate of Elongation
1. RNA polymerase can lay down and connect about 40 nucleotides a second
a) It takes about 25 seconds to transcribe an average length gene of 1000 nucleotides
IV. TERMINATION AND RELEASE
A. Base sequence specified
1. Termination of RNA synthesis occurs at specific base sequences within the DNA
molecule
B. Types of termination sequences
1. Simple (intrinsic) terminators
a) Factor-independent termination sequences
2. Extrinsic Terminators
a) Require termination factors such as rho
b) Rho-dependent termination sequences
V. RNA MOLECULES
A. Major classes of RNA
1. Messenger RNA
a) mRNA
b) An informational molecule used in translation
2. Ribosomal RNA
a) rRNA
b) Structural molecules that forms part of the ribosome
3. Transfer RNA
a) tRNA
b) Both informational and structural
4. Small nuclear RNA (snRNA)
a) Only in eukaryotes
B. Messenger RNA
1. Function
a) The base sequence of DNA determines the amino acid sequence of every
polypeptide chain in the cell
(1) mRNA in eukaryotes deliver this information to the cytoplasm where transcription can occur
b) Its sequence is used to direct amino acid polymerization
(1) Three nucleotides code for one protein
(2) The three nucleotides are collectively referred to as a codon
2. Cistrons
a) Definition
(1) A DNA segment corresponding to one polypeptide chain plus the translational start and stop
signals
b) Monocistronic
(1) A mRNA encoding a single polypeptide
c) Polycistronic
(1) A mRNA encoding several different polypeptide chains
(2) Common in prokaryotes
3. Size
a) Smallest proteins are about 50 amino acids
(1) 150 minimum for a monocistronic mRNA
b) Moderate size polypeptides are 300 – 600 amino acids
(1) 900 – 1800 ribonucleotides
c) Polycistronic mRNA encoding several polypeptides
(1) 3000 – 8000 nucleotides
4. Parts of mRNA
a) Reading frame
(1) The section of mRNA coding for polypeptide amino acid sequence
b) Leader
(1) RNA upstream of the first reading frame
(2) Non-translated
c) Spacer
(1) Between reading frames
(2) Non-translated
d) Tail
(1) Region downstream from the last reading frame
C. Differences between eukaryotes and prokaryotes
1. 5’ and 3’ termini are modified
a) A complex structure called a cap is found at the 5’ end of all mRNA molecules
b) Poly A tail is found on most 3’ ends of mRNA molecules
(1) Up to 20 nucleotides
2. mRNA is processed
a) The primary transcript contains exons and introns
(1) Exons code for amino acid sequences
(2) Introns are intervening sequences that do not code for amino acid sequences
b) Introns are removed from primary transcript
(1) Exons are linked together to form mRNA molecule
3. mRNA is monocistronic
4. Many mRNA molecules are very long-lived