11/17/2015

The Products of Transcription

Fundamental Genetics
Lecture 12

Translation and
Proteins

Messenger RNA (mRNA)

primary structure
linear sequence of RNA bases
carries the genetic information in
the form of codons

Codons

Ribosomal RNA (rRNA)

assumes a 3D structure
(complexed with proteins)
site of protein synthesis

Transfer RNA (tRNA)

John Donnie A. Ramos, Ph.D.
Department of Biological Sciences
College of Science
University of Santo Tomas

assumes a cloverleaf structure

carries amino acids from
cytoplasm to ribosomes

Ribosome

Transfer RNA
Encoded by rDNA
(ribosomal gene)
Synthesized by RNA
polymerase I in the
nucleolus

75-90 nucleotides
Nucleotides are post-transcriptionally
modified
2D cloverleaf structure (Rodbert Holley)
due to base pairing

Complex of RNA and

proteins (monosome)
Prokaryotes: 10K/cell

3D Structure

2D Structure

Steps in Translation

Amino acid binding site - ends

in CCA3 and 5G
Anticodon loop contains
RNA bases complementary to
the codons
Other loops serves as
recognition sites for enzymes
during translation

Initiation of Translation

1. Charging of tRNA
2. Initiation of translation
3. Elongation of polypeptide chain
4. Termination of translation
Charging of tRNA
Loading of specific amino acid to its own tRNA
Catalyzed by aminoacyl tRNA synthetase
32 different tRNA (despite the presence of 61
codons (bec. Of wobbling mechanisms)
20 different aminoacyl tRNA synthetases
Isoaccepting tRNA tRNA that binds to aa
End product: aminoacyl-tRNA complex

Shine-Dalgarmo sequence (5AGGAGG3)

sequence that precedes the first codon
in prokaryote m RNA
Formylmethionine (fmet) the first amino
acid of most polypeptides

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Elongation of Polypeptide Chain

Protein Factors Involved in Translation

Peptidyl site (P site) contains the elongating

peptide
Aminoacyl site (A site) contains the amino acid to
be added
Exit site (E site) exit of uncharged tRNA
Peptidyl transferase catalyzes the formation of
peptide bond
High efficiency (error rate 10-4)
Rate of elongation: 15 aa/sec at 37 C (E. coli)

Termination of Translation
Signaled by stop codons (UAG, UAA,
UGA)
Release Factor 1 (RF1) recognizes stop
codon UAA and UAG
Release Factor 2 (RF2) recognizes stop
codons UGA and UAA
Release factors are GTP dependent
Post-translational modification starts
after release from ribosome

Polyribosomes
Single mRNA being used by different ribosomes for the process of
translation
Also called polysomes
A mechanism to produce more polypeptide (protein) copies

Translation of hemoglobin
mRNA in rabbit reticulocyte

Translation in Eukaryotes
mRNAs stays in the cytoplasm for longer periods before degradation by
RNAses (hours)
Ribosomes are much bigger
mRNA is capped with 7-methyguanosine (7MG)
mRNA contains an initiation sequence called Kozak sequence (ACCAUGG)
discovered by Marilyn Kozak
Formylmethionine (fMet) is not required for initiation but met is often used
as a start codon
More complex protein factors involved in different steps
Elongating polypeptide enters the ER immediately as translation occurs

Translation in giant salivary

gland cells of midgefly

Proteins Form Phenotypes

Phenyketonuria
Mental retardation
Autosomal
recessive
Inability of Phe to
converted to Tyr
Accumulation of
Phe and its
derivatives in
cerobrospinal fluid

Alkaptonuria
Autosomal recessive
Darkening ears and nose
Benign arthritic conditions

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Genes and Proteins

One gene: one enzyme
hypothesis
Proposed by George Beadle
and Edward Tatum (1940s)
Experiments in Neurospora
mutants

Genes and Proteins

One gene: one protein (polypeptide chain)
Not all protein are enzymes
Example: Sickle Cell Anemia (mutant hemoglobin)

Amino Acids

Protein Structure

Primary Structure

Tertiary Structure

Post-translational Modifications
Cleavage of formylmethionine
Cleavage of signal peptides
Acetylation of amino group
Phosphorylation of certain amino acids
Glycosylation
Trimmining of polypetides
Addition of metallic groups
Molecular Chaperons help proteins undergo correct protein folding
to become functional molecules.

Secondary Structure

Quaternary Structure

Protein Function
Structural Function
Collagen
Keratin
Actin
Myosin

Regulatory Function
Hormones
Hemoglobin
Myoglobin

Defense Function
Antibodies
Complement proteins

Catalytic Function
Enzymes
Ribozymes

Others
Histones
Receptors

Enzyme Activity

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Protein Domains and Exon Shuffling

Structural domains of a fibronnectin molecule

DNA organization of an LDL receptor gene