RNA Translation

RNA Translation: RNA makes Protein

In principle:
Translation of messenger RNA (mRNA) takes place on ribosomes,
which include ribosomal RNA (rRNA),
with the help of transfer RNA (tRNA)

Structure of rRNA & tRNA

ribosomal RNA (rRNA)
rRNA + ribosomal protein ribosomes
Structure of rRNA: stems & loops

Structure of eukaryotic ribosome subunits
Large Subunit + Small Subunit = 80S monosome

A site (Aminoacyl), P site (Peptidyl), & E site (Exit) [APE or EPA complex]

transfer RNA (tRNA)
       adaptor molecule: ~30 tRNA types
       2-dimensional 'cloverleaf' model
            small: 75 ~ 90 nucs
       stems & loops
            D-loop & TC-loop ( = pseudo-uridylic acid)
                   tRNA characterized by 2^omodified bases
            amino-acceptor stem
                    3' - ~~~~ CACCA - 3'
                    5' -~~~~ G    - 5'
            anticodon loop
                   specificity of tRNA for mRNA determined by 3-ribonucleotide sequence

3-dimensional structure an "L"
D- & TC-loops fold back on each other

Charged tRNA: aminoacyl synthetase_(x) forms ester linkage between
       3'-A of amino-acceptor stem of tRNA_(x)
       & COOH of amino acid_(x)
       ~20 synthetase types 'recognize' correct anticodon loop
       isoacceptance:
           one-to-one correspondence between synthetase & amino acid

RNA Translation: Protein Synthesis

Ribosomes "read" mRNA & assemble polypeptide according to the Genetic Code

Initiation at start codon (AUG)
SSU binds at Shine-Delgarno sequence (-6 nucs)
Multiple complexes form on a single mRNA: polysome (polyribosome)

tRNA_met always added first [N-formyl-methionine in prokaryotes]

In simplified form

Elongation: addition of amino acids according to Genetic Code

       P site amino acid transferred to A site amino acid
              uncharged tRNA released from P site, passes to E site
amino end of initial met remains unchanged
   and so on ...

   "Wobble": pairing of codon / anticodon goes 5'3' on codon
                      last position can miss-pair with either purine / pyrimidine
                            Fewer tRNA species needed:
                       Ex.: three tRNA_ser species for six codons

Termination: release of polypeptide

      stop codon (UAG, UAA, or UGA) enters A site
                release factor cleaves polypeptide from terminal tRNA_n
                polypeptide product: C - lys - pro - gly - phe - met - N

Griffiths et al. (1996) Fig. 13-7 is a nice schematic summary (HOMEWORK #11)

Bioinformatics of DNA, mRNA, & Protein

This is a logical, not a biochemical, relationship:
        Because mRNA is transcribed from the template strand,
        it "looks like" the sense strand (except for 'U').
        information content of the DNA sense strand and mRNA are identical

Protein sequences can be read directly from DNA:
         Read the sense strand in the 5'3' direction,
         Substitute 'T' for 'U' in the code table [or in your head]
       Computer programs ( MEGA, etc.) do this automatically

There are three reading frames on either strand
X two 5'3' strands six possible ways to read dsDNA
Open Reading Frames suggest protein sequences

Deducing protein sequences from "shotgun" DNA sequences: a major research activity
Bioinformatics: extraction of information from large macromolecular data sets

These clues are useful:
         Remember that all coding sequences:
            are read only in the 5'3' direction
            begin with a "start" (AUG) codon
          end with a "stop" (UAG, UAA, or UGA) codon.
             Ex.: a typical exam problem is to identify a polypeptide of six amino acids from a dsDNA molecule

          But: in real life research, any large sequence of eukaryotic DNA
                may not have start and (or) stop codons for complete protein,
                   [and most AUG codons are not 'start' codons]
                and may be include an intron with one or more 'stop' triplets .

Do not assume that dsDNA molecules read left to right, on top strand

Homework #12:

Practice DNA "Translation" problems; there's an App for that: RandORF