From Gene to Protein: Chpt. 17

How does the DNA lead to specific traits??? Archibald Garrod (1909) hypothesized: “proteins are the link between genotype & phenotype”

1909 - Archibald Garrod Suggested genes control enzymes, & enzymes catalyze chemical processes in cells. Inherited Diseases are “inborn errors of metabolism” where a person can’t make an enzyme.”

Example Alkaptonuria (Al ka toe nuria)- where urine turns black after exposure to air b/c of chemical alkapton (Al kae ton). these individuals must lack an enzyme to metabolize (break down) alkapton (the chemical).

enzyme enzyme enzyme enzyme

George Beadle American(1930) *“ mutations in eye color (Drosophila) are a result of blocks in pigment production” *“no enzyme is made - therefore pathway to produce color pigment is not complete… white eyes” Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from

G. Beadle & Edward Tatum Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from Beadle and Tatum set out to provide experimental proof of the connection between genes and enzymes.

Nobel Prize in Physiology or Medicine 1958 G. Beadle & Edward Tatum Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from Cal Tech. Rockefeller Inst. NY, NY Nobel Prize in Physiology or Medicine 1958 "for their discovery that genes act by regulating definite chemical events"

G. Beadle & Edward Tatum test organism = Neurospora Neurospora (pink bread mold) Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from

G. Beadle & Edward Tatum mutant molds (not wild type) had a variety of special nutritional needs. Unlike their wild type counterparts, they could not live without the addition of particular vitamins or amino acids to their food. Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from

*“ various mutations must be G. Beadle & Edward Tatum *“ various mutations must be abnormal variations of genes” Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from

wild type mold survived in minimal medium agar. One Gene, One Enzyme Hypothesis -Beadle & Tatum this is the normal metabolic pathway wild type mold survived in minimal medium agar. The wild type mold can produce all of the enzymes it needs to produce the necessary amino acids to live. Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from

Mutants did not survive in minimal medium agar. One Gene, One Enzyme Hypothesis -Beadle & Tatum Mutants did not survive in minimal medium agar. The mutants must not produce all of the enzymes needed Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from The enzymes would synthesize particular amino acids. But they could survive in complete agar… supplemented with all 20 amino acids.

One Gene, One Enzyme Hypothesis -Beadle & Tatum Experiment Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from

One Gene, One Enzyme Hypothesis -Beadle & Tatum Tried to identify where the mutants’ metabolic defects (mutations) were Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from

One Gene, One Enzyme Hypothesis -Beadle & Tatum All strains were grown on complete media. This contained all a.a. & vitamins made by the wild type, they all thrived… Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from By the way, this mold MUST make Arginine in order to grow.

One Gene, One Enzyme Hypothesis -Beadle & Tatum Took mutants out of the complete media They did not all grow when put on “minimal media” Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from They figured that mutants were unable to produce compounds essential for growth… perhaps their genes did not code for an enzyme… this ultimately did not let the organism produce arginine??

One Gene, One Enzyme Hypothesis -Beadle & Tatum Put each mutant type in different vials containing minimal + one amino acid. (in this pix., the a.a. is ornathine) Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from

Because Class I grew on minimal medium supplemented with ornithine, citrulline, or argine, it had to be missing enzyme A. This would be required to form all three compounds Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from

One Gene, One Enzyme Hypothesis -Beadle & Tatum Class II could not produce citruline, and that is needed for the rest of the pathway to occur... Ornathine “piled up” Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from

One Gene, One Enzyme Hypothesis -Beadle & Tatum The media that allowed growth, would show where the metabolic defect (mutation) occurred there was always a “pile up” of the amino acid before Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from

One Gene, One Enzyme Hypothesis -Beadle & Tatum If we inactivate one gene, that would code for an enzyme to carry out X -> Y conversion, no product X or Y is produced. If product Y is needed to ultimately produce arginine, no growth will occur Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from

One Gene, One Enzyme Hypothesis -Beadle & Tatum The supplement that allowed growth, would show where the metabolic defect (mutation) occurred Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from

One Gene, One Enzyme Hypothesis -Beadle & Tatum Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from

One Gene, One Enzyme Hypothesis -Beadle & Tatum ex. this mutant grew (was able to complete the pathway) in the vial supplemented with Citrulline. Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from therefore, it must be defective in synthesizing Citrulline~ornithine piled up

each gene dictates the production of one enzyme One Gene, One Enzyme Hypothesis -Beadle & Tatum each gene dictates the production of one enzyme each mutant lacks the ability to produce an enzyme Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from

One Gene, One Polypeptide Hypothesis -Beadle & Tatum (revisited) *Some proteins are not enzymes (ex. insulin) *Some proteins are made of more than one polypeptide chain (hemoglobin) *Each chain specified by its own gene Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from One Gene, One Polypeptide Hypothesis -Beadle & Tatum (revisited)

How are enzymes (proteins/polypeptides) made? Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from

How do we get from DNA to protein? Beadle and Tatum Found that certain enzymes were missing the enzymes to synthesize particular amino acids As long as the amino acids were provided for them they could grow but they could not grow on a minimal nutritional medium These mutants called auxotrophs can grow if all 20 amino acids are provided but the must have increased nourishment which is where their name comes from

RNA

RNA is the bridge between DNA and Protein mRNA The genetic message is read based on triplet coding patterns. The triplet code is read by RNA (codon) and is then translated to another type of RNA (anticodon) which turns the language from nucleic acid to amino acid tRNA

This is the basic process: TRANSCRIPTION TRANSLATION The genetic message is read based on triplet coding patterns. The triplet code is read by RNA (codon) and is then translated to another type of RNA (anticodon) which turns the language from nucleic acid to amino acid

No nucleus, protein synthesis occurs same time as transcription. Transcription occurs in nucleus-> pre -mRNA Translation occurs on ribos.

DNA --> RNA --> PROTEIN!

Triplets - series of three nucleotides- code for Amino Acids U not T

(1961) Marshall Nirenberg deciphered first codon.

(1961) Marshall Nirenberg deciphered first codon. Won Nobel prize in Physiology and Medicine 1968 for the interpretation of the genetic code and its function in protein synthesis.

1965 Dictionary of Amino Acids This is the same for ALL species of organisms RNA also reads in the 5’ to 3’ direction . This is an amino acid map which tells the amino acid based upon which nucleotides are transcribed. There are 64 possible codons 61 of which code for a specific amino acid, the others are stop (2) and Start(1)

1965 Dictionary of Amino Acids Evolutionary Significance?? RNA also reads in the 5’ to 3’ direction . This is an amino acid map which tells the amino acid based upon which nucleotides are transcribed. There are 64 possible codons 61 of which code for a specific amino acid, the others are stop (2) and Start(1)

Code Redundancy Third base in a codon shows "wobble”. First two bases are the most important in reading the code and giving the correct AA. The third base often doesn’t matter.