How does the cell manufacture these magnificent machines? Proteins, that is…

Single-letter code: M D L Y

Cys - S - H + H - S - Cys Cys - S - S - Cys Disulfide bridge formation stabilizes protein structure

Proteins are 3-dimensional molecules Primary structure = Amino acid sequence Secondary structure = 1.Alpha helix 2.Beta sheet Tertiary structure = 3-D shape Quaternary structure = ??  -helix  -sheet

The Genetic Code How the genetic code was deduced is quite an interesting but horribly complicated story of prokaryotic genetics. I’ll just give you the Cliff notes version: Francis Crick and Sidney Brenner figured out that: The genetic code maps ‘codons’ of 3 bases into one amino acid. AUA->Ile GAU->Asp AGA->Arg mRNA-> Amino Acid

The Genetic Code Crick and Brenner figured out that: The DNA code is read sequentially from a fixed position in the gene

The mechanism and machinery for translating a protein Three components:

tRNA is the adapter. Complementary to the Codon Amino acid is matched to the Anti-codon.

Translation occurs 5’ to 3’ The translation is performed with the help of the Ribosome.

RNA is the major component of the Ribosome. About 2/3 of the Ribosome is RNA by mass. These RNA molecules are called rRNA and they play a central role in the translation of mRNA into polypeptides.

mRNA, rRNA, tRNA and protein synthesis In translation, the language of nucleic acids is translated into a new language, that of proteins mRNA provides the code, in linear digital form, for making a protein tRNA provides an adaptor that links the code in a polynucleotide chain to amino acids that make up the polypeptide chain rRNA and ribosomes provide the decoder. Ribosomes bring together mRNA and tRNA, and catalyze the translation of an mRNA into a polypeptide chain. Ribosomes are the site of protein synthesis. Ribosomes create peptide bonds between amino acids to create proteins

Translation normally occurs on polyribosomes, or polysomes This allows for amplification of the signal from DNA and RNA, i.e.,… One gene copy Hundreds of mRNAs Thousands of proteins

rRNAs form complex 2 o and 3 o structures that are essential for their function What does this rRNA molecule look like in three dimensions?

3-D model of 16s rRNA molecule as it folds in the ribosome… …and overlaid with its protein subunits

Behold, the ribosome! Proteins are blue, RNAs are red and white

Two views of the adaptor molecule, transfer RNA (tRNA), which guides amino acids to the mRNA-ribosome complex The anticodon of the tRNA aligns with the codon in mRNA through complementary base pairing

A E Large subunit P Small subunit Translation - Initiation fMet UAC GAG...CU-AUG--UUC--CUU--AGU--GGU--AGA--GCU--GUA--UGA-AT GCA...TAAAAAA 5’ mRNA 3’

A E Ribosome P UCU Arg Aminoacyl tRNA Phe Leu Met Ser Gly Polypeptide CCA Translation - Elongation GAG...CU-AUG--UUC--CUU--AGU--GGU--AGA--GCU--GUA--UGA-AT GCA...TAAAAAA 5’ mRNA 3’

A E Ribosome P Phe Leu Met Ser Gly Polypeptide Arg Aminoacyl tRNA UCUCCA Translation - Elongation GAG...CU-AUG--UUC--CUU--AGU--GGU--AGA--GCU--GUA--UGA-AT GCA...TAAAAAA 5’ mRNA 3’

A E Ribosome P CCA Arg UCU Phe Leu Met Ser Gly Polypeptide Translation - Elongation GAG...CU-AUG--UUC--CUU--AGU--GGU--AGA--GCU--GUA--UGA-AT GCA...TAAAAAA 5’ mRNA 3’

A E Ribosome P Translation - Elongation Aminoacyl tRNA CGA Ala CCA Arg UCU Phe Leu Met Ser Gly Polypeptide GAG...CU-AUG--UUC--CUU--AGU--GGU--AGA--GCU--GUA--UGA-AT GCA...TAAAAAA 5’ mRNA 3’

A E Ribosome P Translation - Elongation CCA Arg UCU Phe Leu Met Ser Gly Polypeptide CGA Ala GAG...CU-AUG--UUC--CUU--AGU--GGU--AGA--GCU--GUA--UGA-AT GCA...TAAAAAA 5’ mRNA 3’

A E Ribosome P Translation - Termination CGA Phe Leu Met Ser Gly Polypeptide Ala Arg Val CGA GAG...CU-AUG--UUC--CUU--AGU--GGU--AGA--GCU--GUA--UGA-AT GCA...TAAAAAA 5’ mRNA 3’ STOP

A E P Translation - Termination CGA Phe Leu Met Ser Gly Polypeptide Ala Arg Val CGA GAG...CU-AUG--UUC--CUU--AGU--GGU--AGA--GCU--GUA--UGA-AT GCA...TAAAAAA 5’ mRNA 3’ STOP

How many bases are required to make a genetic code to serve 20 different amino acids? # of 2-base combos = 4 2 = 16 # of 3-base combos = 4 3 = 64 Not enough! Too many! What is the solution? Evolve a code that is redundant! How? Degeneracy at the third codon position Let’s look at the Genetic Code

Transfer RNA (tRNA) and the genetic code

Many different base modifications occur in tRNA Why are these modifications necessary?

Transfer RNA (tRNA) and the genetic code Legitimate G-U bp Wobble bp

Point mutations Point mutations can affect protein structure and function Type of point mutations: - substitutions (missense and nonsense mutations) - insertions and deletions (frameshift mutations)

Frame Shift Mutations What happens when you get insertions or deletions of bases in the DNA sequence? Usually you end up with a mess. THE BIG FAT CAT ATE THE RAT AND GOT ILL Deletion of one base THE IGF ATC ATA TET HER ATA NDG OTI LL And its all pops and buzzes. Usually frame shift mutations result in premature stop codons.

Where can you get more information about the basic concepts embedded within the Central Dogma of Molecular Biology? Here is a great site, full of simple, clear, and animated (!) tutorials: Chapters in this series provides an excellent review of the first six lectures in Bioinformatics:

Ribosomes are large ribonucleoprotein (RNP) complexes They are complex affairs, composed of an array of RNA + proteins