Ch. 17: From Gene to Protein

The Connection Between Genes and Proteins

• The study of metabolic defects provided evidence that genes specify proteins– Garrod, suggested phenotypes had to do with expression of

genes for enzymes• Transcription and translation are the two main processes

linking gene to protein– Copy the information and interpret the information

• In the genetic code, nucleotide triplets specify amino acids– Sequence of nucleotides = primary protein structure

• The genetic code must have evolved very early in the history of life– DNA code is universal…..all cells use the same codons and

amino acids to make their various proteins

Back to Mendel

• One of Mendels “factors” for peas was stem length. We say “height” and the alleles are tall and short. Actually it’s “length” and the stems are long or not-long

• Normal peas have a gene for the hormone called gibberellin which stimulates stem elongation.

• Dwarf peas lack this gene and do not make gibberellin and are therefore not tall.

• Dwarf peas will grow to normal height if gibberellins are added to their water

• PROTEINS ARE THE LINKS BETWEEN GENOTYPE AND PHENOTYPE.

Scientific Evidence

• 1909 – “inborn errors of metabolism” – alkaptonuria

• 1930 – Beadle and Ephrussi, eye color in flies is due to an enzyme for pigment production

• Beadle and Tatum – minimal medium Neurospora crassa (bread mold), used x-rays to create mutations, complete media had 20 amino acids, looking for inability to metabolize amino acids from a limited source– Mutants had defects in metabolism– Must be enzymes related– Enzymes are proteins

– One gene – one enzyme hypothesis– now modified to one gene – one (protein) polypeptide

Beadle and Tatum’s Neurospora crassa Experiment

Transcription and Translation

• Genes have instructions for making proteins, but genes do not make proteins directly

• Transcription is the synthesis of RNA under the direction of DNA. DNA provides the template. Get an accurate copy; mRNA

• Translation is the actual synthesis of a polypeptide, at the ribosome, under the direction of the mRNA

• DNA RNA protein (polypeptide)

Terminology

• Triplet code: three DNA nucleotides = a word• mRNA: carries message from DNA to ribsome• tRNA: transports amino acids within cytoplasm

rRNA: ribosomes are composed of rRNA and proteins• Ribosome: solid organelle found in cytoplasm of ALL cells;

used to manufacture protein• Template strand: for each gene only one side of the DNA is

transcribed• Codon: mRNA triplets are called codons• Reading frame: 5’3’, starting at beginning, groups of three

– The red dog ate the cat– xHer edd oga tat hec atx

Cracking the Code

• 1960 – Marshall Nirenberg at NIH– National Institute of Health– www.nih.gov– Human genome projects library

• Translated all the possible codons into amino acids

• Found codons for “start” and for “stop” • Several amino acids can be coded for with more

than one codon (redundancy) but no codons are for multiple amino acids (no ambiguity)

• “wobble effect”

Code Evolved Early in the History of Life on Earth (and any life anywhere else too)

• Code is (near) universal to all know/studied organisms…. Bacteria can translate human genetic information

• All modern organisms have a common ancestor

• Few exceptions are found in protista and in mitochondria DNA (…. Endosymbiont hypothesis….)

• Sketch a DNA molecule with chemically correct details

• Show how it would replicate and • How it would transcribe and• List the amino acids in the short polypeptide it

forms• It has the following template strand sequence of

DNA triplets– TAC TTT GAG ATT

Genomic like information

• Stthegeneticcodeisnearlyuniversalpsharedbyorganismsfromthesimplestbacteriatothemostcomplexplantsandanimalspstthernacodonccgpforinstancepistranslatedastheaminoacidprolineinallorganismswhosegeneticcodehasbeenexaminedpstinlaboratoryexperientspgenescanbetranscribedandtranslatedaftertheyaretransplantedfromonespeciestoanotherpfponeimportantapplicationisthatbacteriacanbeprogrammedbytheinsertionofhumangenestosynthesizecertainhumanproteinsthathaveimportantmedicalusesp

The Synthesis and Processing of RNA

• Transcription is the DNA directed synthesis of RNA

• Eukaryotic cells modify RNA after transcription

Transcription

• RNA polymerase fits onto DNA (3’) and moves in a 5’ 3’ direction for the synthesis of the RNA strand.

• C with G and this time, A with U (uracil)• DNA acts as a template • DNA is only opened at a small region (gene or

genes of interest)• DNA helix reseals as RNA polymerase passes

by…. Completely intact and undiluted.

Bacterial transcription

• Eukaryotic cells have 3 kinds of RNA polymerase (I, II – used in RNA synthesis and III)

• Bacteria have one kind – it makes not only mRNA but also other types of RNA

• Bacteria have one chromosome and many plasmids. Information is constantly being sent to ribosomes for translation into proteins needed by the bacterial cell

Steps of Transcription1. Initiation

Promoter – region where polymerase attaches and a dozen bases upstream; start here and use this side of the helix.

Collection of transcription factors initiate the “complex” – TATA box2. Elongation

DNA exposed 20 bases at a time5’ 3’ synthesis of RNA strandRNA peels away from DNA as completedrate is 60 nucleotides per second

3. TerminationDNA contains a terminator sequencepolymerase continues to a AAUAAA sequence and 10-35 nucleotides later the preRNA is cut freeother details are still ‘murky’

Modification of RNA

• Initially RNA is called preRNA• The 5’ end (transcribed 1st) is capped with

special guanine – provides protection and a start here signal for translation

• Other end gets a ploy A tail (AAA-AAA) – in addition to ribosomal attachment and protection, it seems to facilitate RNA as it leaves the nucleus

• These regions are nontranslated

Further modification of RNA

• Most of the pre RNA is actually removed…. It didn’t code for information about how to make a protein. We are uncertain of the function of this info, which does not make the info unimportant.

• Initially the RNA can be 8000 bases, actual info for protein that goes to ribosomes is about 1200 or 400 amino acids (1200 bases/ 3 bases per codon)

“Cut and Paste”• Called RNA splicing• Introns (intervening segments) are removed

– they are noncoding, short, repetitive sequences, unique – cause restriction enzymes to cut segments differently and create the DNA fingerprint

– Probably have a role in gene expression and activity– May be place where new proteins evolve– Increase odds of crossing over during synapsis of tetrads (meiosis II)

• Exons (expressed) – these are translated into amino acids for the polypeptide – 150 nucleotides

• 5’ cap + exon + exon + exon + …. + poly A tail• Process requires snRNP’s - small nucleotide ribonucleoproteins….

Sites to bind• Ribozymes = RNA that functions as an enzyme.