CHAPTER 12 From DNA to Protein: Genotype to Phenotype

Genotype to Phenotype Genes are made up of DNA (genotype). Genes cannot directly produce a phenotype. Genes must be expressed (phenotype) as polypeptides.

DNA, RNA, and the Flow of Information RNA differs from DNA in three ways: It is single-stranded, Its sugar molecule is ribose rather than deoxyribose, and Its fourth base is uracil rather than thymine. Adenine pairs with uracil

DNA, RNA, and the Flow of Information The central dogma of molecular biology is DNA  RNA  protein. Review Figure 12.2

DNA, RNA, and the Flow of Information - Summary A gene is transcribed to produce messenger RNA (mRNA). mRNA is complementary to one of the DNA strands Transfer RNA ((tRNA) translates sequence of bases in mRNA into appropriate sequence of amino acids. Amino acids join together (peptide bonds) to form proteins. Review Figure 12.3

figure 12-03.jpg Figure 12.3 Figure 12.3

Transcription – synthesis of RNA from DNA Transciption requires the enzyme RNA polymerase, RNA nucleotides, and a DNA template. Transcription occurs in the nucleus. The product, a RNA transcript, is sent to the cytoplasm where translation occurs.

Transcription Transcription is divided into three processes: Initiation, Elongation, and termination

Initiation Transcription begins at a promoter – a special sequence of DNA . The promoter determines the direction, which strand to read, and direction to take RNA polymerase binds to the promoter. Once the polymerase is attached to the promoter DNA, the DNA strands unwind and transcription begins.

Elongation As RNA polymerase moves along the DNA, it continues to unwind DNA about 20 bases pairs at a time. One side of the unwound DNA acts as template for RNA synthesis RNA transcript is formed by complementary base pairings.

Termination Specific DNA base sequences terminate transcription. Pre-mRNA is released. Review Figure 12.4 The resulting RNA transcript may be mRNA, tRNA, or rRNA.

Messenger RNA (mRNA) carries a genetic message from DNA to the protein synthesizing machinery of the cell (ribosomes)

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The Genetic Code The genetic code consists of triplets of nucleotides (codons). Since there are four bases, there are 64 possible codons (43) There are more codons than different amino acids.

The Genetic Code AUG codes for methionine and is the start codon. UAA, UAG, and UGA are stop codons. Stop codons indicate the end of translation. The other 60 codons code only for particular amino acids.

The Genetic Code Since there are only 20 different amino acids, the genetic code is redundant; that is, there is more than one codon for certain amino acids. However, a single codon does not specify more than one amino acid. Review Figure 12.5

The Universal Genetic Code The genetic code appears to be nearly universal. Provides a common language for evolution. Implications for genetic engineering.

figure 12-05.jpg Figure 12.5 Figure 12.5

Preparation for Translation: Linking RNA’s, Amino Acids, and Ribosomes Translation occurs at the ribosomes. Translate the message from sequence of nucleotides to sequence of amino acids

Components of Translation Ribosomes – small and large subunits mRNA (messenger RNA) tRNA (transfer RNA) tRNA transfer an amino acid. tRNA has a sequence of 3 bases known as the anticodon that is complementary to mRNA codon amino acids are linked in codon-specified order per mRNA.

figure 12-07.jpg Figure 12.7 Figure 12.7

Example of Process The DNA coding region for proline is GGG which is transcribed to The mRNA codon CCC which binds to The tRNA with the anticodon GGG

Activating Enzymes link tRNA and amino acids A family of activating enzymes – aminoacyl-tRNA synthetases_ attach specific amino acids to their appropriate tRNA’s to from charged tRNA The amino acid is attached to the 3’ end of tRNA with a high energy bond Review Figure 12.8

Ribosomes The ribosome consist of a large and a small subunit. When not active in translation, the ribosomes exist as separate units. They can come together and separate as needed. Review Figure 12.9

figure 12-09.jpg Figure 12.9 Figure 12.9

Three Phases of Translation Initiation Elongation Termination

Translation: Initiation A sequence of mRNA (initiation factors) binds to the small subunit of a ribosome. Aminoacyl tRNA bearing UAC binds to the start codon. Large subunit of ribosome joins the complex Review Figure 12.10

figure 12-10.jpg Figure 12.10 Figure 12.10

Elongation – A Four Step Process A charged tRN moves into the ribosome and occupies the A site. Its anticodon matches the mRNA codon The polypeptide chain is transferred. Ribosome moves along the mRNA. Empty tRNA is ejected via the E site. tRNA with peptide chains moves to P site. A is empty. Repeat

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Translation: Termination The presence of a stop codon (UAA, UAG, or UGA) in the A site of the ribosome causes translation to terminate. Both tRNA and the polypeptide are released from the P site. The ribosomes separate Review Figure 12.12

figure 12-12.jpg Figure 12.12 Figure 12.12

4 Sites for tRNA Binding T (transfer) site is where tRNA + amino acids first attaches to the ribosome. The A (amino acid) site is there the tRNA anticodon binds to mRNA codon The P (polypetide) site is where the amino acids are bonded together. The E (exit) site is where the tRNA will leave the ribosome to pick up additional amino acids.

Regulation of Translation Antibiotics can interfere with translation Erythromycin plugs the exit channel so the polypeptide chain cannot leave the ribosome. Review Table 12.2

Regulation of Translation In a polysome, more than one ribosome moves along the mRNA at one time. Multiple copies of the same protein is made for a single mRNA. Review Figure 12.13

figure 12-13.jpg Figure 12.13 Figure 12.13

Posttranslational Events The functional protein may vary from the polypeptide chain that is originally released. Signals contained in the amino acid sequences of proteins direct them to cellular destinations. And polypeptides may be altered by the addition of chemical groups that affect function of the protein. Review Figure 12.14

figure 12-14.jpg Figure 12.14 Figure 12.14

Posttranslational Events Protein synthesis begins on free ribosomes in the cytoplasm. Those proteins destined for the nucleus, mitochondria, and plastids are completed in the cytoplasm and have signals that allow them to bind to and enter destined organelles.

Posttranslational Events Proteins destined for the ER, Golgi apparatus, lysosomes, and outside the cell complete their synthesis on the ER surface. They enter the ER by the interaction of a hydrophobic signal sequence with a channel in the membrane. Review Figure 12.15

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Posttranslational Events Covalent modifications of proteins after translation include: proteolysis – polypeptide chain is cut Glycosylation – additions of sugars to proteins Phosphorylation – add phosphate groups to protiens. Review Figure 12.16

figure 12-16.jpg Figure 12.16 Figure 12.16

Mutations: Heritable Changes in Genes Mutations in DNA are often expressed as abnormal proteins. However, the result may not be easily observable phenotypic changes. Some mutations appear only under certain conditions, such as exposure to a certain environmental agent or condition.

Mutations: Heritable Changes in Genes Point mutations (silent, missense, nonsense, or frame-shift) result from alterations in single base pairs of DNA.

Mutations: Heritable Changes in Genes Chromosomal mutations (deletions, duplications, inversions, or translocations) involve large regions of a chromosome. Review Figure 12.18

figure 12-18.jpg Figure 12.18 Figure 12.18

Mutations: Heritable Changes in Genes Mutations can be spontaneous or induced. Spontaneous mutations occur because of instabilities in DNA or chromosomes. Induced mutations occur when an outside agent damages DNA. Review Figure 12.19

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One Gene, One Polypeptide Certain hereditary diseases in humans have been found to be caused by the absence of certain enzymes. Table 3-4 Phenylketonuria (PKU) is a recessive disease caused by a defective allele for phenylalanine hydroxylase. In the absence of the enzyme, phenylalanine in food is not broken down and accumulates.

PKU At high concentrations phenylalanine is converted to phenylpyruvic acid. Phenylpyyruvic acid interferes with the development of the nervous system. Solution: An infant is put on a low phenylalanine diet so phenylalanine does not accumulate, no phenylpyruvic acid is made and the child develops normally. Figure 3-28 These observations supported the one-gene, one-polypeptide hypothesis.