Chapter 14 From DNA to Protein

Objectives 1. Know how the structure and behavior of DNA determines the structure and behavior of the three forms of RNA during transcription. 2. Understand the role the three forms of RNA play in determining the primary structure of polypeptide chains during translation. 3. Understand the types of mutations and their role in genetic variation.

14.0: Ricin and Your Ribosomes Ricin – comes from the castor oil plant, and can be used as a bioweapon. Ricin has deadly effects – it inactivates ribosomes, the protein-building machinery of all cells. Protein synthesis stops, cells die, the individual dies, there is no antidote.

Life depends on enzymes Enzymes are proteins. All proteins consist of polypeptide chains. The chains are sequences of amino acids that correspond to genes – sequences of nucleotide bases in DNA. Enzymes may selectively unwind certain regions of DNA to encode information to make proteins.

The path from genes to proteins DNA RNA Protein The two steps include: Transcription Translation

Steps from DNA to Proteins Same two steps produce all proteins: 1) DNA is transcribed to form RNA Occurs in the nucleus RNA moves into cytoplasm 2) RNA is translated to form polypeptide chains, which fold to form proteins

14.1 How is RNA Transcribed from DNA The first step in protein synthesis, is transcription, a sequence of nucleotide bases is exposed in an unwound region of a DNA strand. That sequence is the template upon which a single strand of RNA is assembled from adenine, cytosine, guanine, and uracil subunits.

Three Classes of RNAs Messenger RNA (mRNA) Ribosomal RNA (rRNA) Carries protein-building instruction or code Ribosomal RNA (rRNA) Combines with proteins to form ribosomes Transfer RNA (tRNA) Delivers the correct amino acids to ribosomes

RNA molecule One phosphate group A five carbon sugar – ribose Four bases: Adenine Cytosine Guanine Uracil – instead of Thymine

A Nucleotide Subunit of RNA uracil (base) phosphate group sugar (ribose) Fig. 14-2, p. 220

Base Pairing during Transcription DNA base pairing during transcription RNA DNA base pairing during DNA replication DNA Fig. 14-2c, p.220

Transcription & DNA Replication Like DNA replication Nucleotides added in 5’ to 3’ direction Unlike DNA replication Only small stretch of DNA is template RNA polymerase catalyzes nucleotide addition Product is a single strand of RNA

Promoter The promoter is a base sequence in the DNA that signals the start of a gene. For transcription to occur, RNA polymerase must first bind to a promoter, and then move along to the end of the gene.

Promoter promoter region RNA polymerase, the enzyme that catalyzes transcription a RNA polymerase initiates transcription at a promoter region in DNA. It recognizes a base sequence located next to the promoter as a template. It will link the nucleotides adenine, cytosine, guanine, and uracil into a strand of RNA, in the order specified by DNA. Fig. 14-3a, p.220

Gene Transcription DNA template at selected transcription site newly forming RNA transcript DNA template winding up DNA template unwinding b All through transcription, the DNA double helix becomes unwound in front of the RNA polymerase. Short lengths of the newly forming RNA strand briefly wind up with its DNA template strand. New stretches of RNA unwind from the template (and the two DNA strands wind up again). Fig. 14-3b, p.220

Adding Nucleotides direction of transcription 3´ 5´ 5´ 3´ growing RNA transcript c What happened at the assembly site? RNA polymerase catalyzed the assembly of ribonucleotides, one after another, into an RNA strand, using exposed bases on the DNA as a template. Many other proteins assist this process. Fig. 14-3c, p.221

Transcript Modification of mRNA unit of transcription in a DNA strand exon intron exon intron exon transcription into pre-mRNA 5’ end Cap 3’ end poly-A tail snipped out non- coding introns snipped out non-coding introns mature mRNA transcript Fig. 14-4, p.221

Modifications of mRNA In nucleus 5’ end is capped with guanine – “start” signal for translations. Cap will also help bind the mRNA to a ribosome 3’ end a “poly-A-tail” is added. 100 – 200 molecules of adenine is added Introns (non-coding) are snipped out, exons (coding portions) are spliced together Alternative splicing results in different proteins

14.2 Genetic Code Set of 64 base triplets Codons 61 specify amino acids 3 stop translation Fig. 14-6, p.222

The Genetic Code Every three bases in mRNA (triplet) code of an amino acid. Both DNA and its RNA transcript are linear sequences of nucleotides carrying the hereditary code. The genetic code consists of 61 triplets that specify amino acids, AUG – “start” codon Methionine, and three “stops”

Genetic Code DNA mRNA mRNA codons amino acids threonine proline glutamate glutamate lysine Fig. 14-5, p.222

14.3 The other RNA’s - tRNA codon in mRNA anticodon amino-acid attachment site amino acid OH Figure 14.7 Page 223

tRNA Each kind of tRNA has an anticodon that is complementary to an mRNA codon Each tRNA also carries one specific amino acid. After the mRNA arrives in the cytoplasm an anticodon on a tRNA bonds to the codon on the mRNA, and thus a correct amino acid is brought into place.

Ribosomes A ribosome has two subunits Each ribosome is composed of rRNA and structural proteins) that come together only during translation. Polypeptide chains are built on ribosomes.

small ribosomal subunit large ribosomal subunit Ribosomes funnel small ribosomal subunit + large ribosomal subunit intact ribosome Fig. 14-8, p.223

14.4 Three Stages of Translation Initiation Elongation Termination

Initiation Initiator tRNA binds to small ribosomal subunit Small subunit/tRNA complex attaches to mRNA and moves along it to an AUG “start” codon Large ribosomal subunit joins complex

Binding Sites binding site for mRNA A (second binding site for tRNA) P (first binding site for tRNA)

Elongation mRNA passes through ribosomal subunits tRNAs deliver amino acids to the ribosomal binding site in the order specified by the mRNA Peptide bonds form between the amino acids and the polypeptide chain grows

Elongation

Termination Stop codon into place No tRNA with anticodon Release factors bind to the ribosome mRNA and polypeptide are released into the cytoplasm. mRNA new polypeptide chain

What Happens to the New Polypeptides? Some just enter the cytoplasm Many enter the endoplasmic reticulum and move through the cytomembrane system where they are modified Polysome- several ribosomes moving along mRNA at the same time.

Overview Transcription Translation mRNA rRNA tRNA Mature mRNA transcripts ribosomal subunits mature tRNA Translation

Fig. 14-9a-e, p.224 elongation binding site for mRNA P (first binding site for tRNA) A (second binding site for tRNA) Amino Acid 1 Amino Acid 1 c Initiation ends when a large and small ribosomal subunit converge and bind together. Amino Acid 2 Amino Acid 2 d The initiator tRNA binds to the ribosome. e One of the rRNA molecules b Initiation, the first stage of translating mRNA, will start when an initiator tRNA binds to a small ribosomal subunit. initiation a A mature mRNA transcript leaves the nucleus through a pore in the nuclear envelope. Fig. 14-9a-e, p.224

Fig. 14-9f-k, p.224 g A third tRNA binds with the next codon f The first tRNA is released h Steps f and g are repeated termination i A STOP codon moves into the area where the chain is being built. j The new polypeptide chain is released from the ribosome. k The two ribosomal subunits now separate, also. Fig. 14-9f-k, p.224

Base-Pair Substitutions 14.5 Gene Mutations A gene mutation is a change in one to several bases in the nucleotide sequence of DNA, which can result in a change in the protein synthesized. Base-Pair Substitutions Insertions Deletions

Base-Pair Substitution a base substitution within the triplet (red) original base triplet in a DNA strand During replication, proofreading enzymes make a substitution possible outcomes: or original, unmutated sequence a gene mutation

Frameshift Mutations Insertion Deletion Both shift the reading frame Extra base added into gene region Deletion Base removed from gene region Both shift the reading frame Result in many wrong amino acids

Frameshift Mutation Fig. 14-10, p.226 part of DNA template mRNA transcribed from DNA resulting amino acid sequence THREONINE PROLINE GLUTAMATE GLUTAMATE LYSINE base substitution in DNA altered mRNA altered amino acid sequence THREONINE PROLINE VALINE GLUTAMATE LYSINE deletion in DNA altered mRNA THREONINE PROLINE GLYCINE ARGININE altered amino acid sequence Fig. 14-10, p.226

Transposons DNA segments that move spontaneously about the genome When they insert into a gene region, they usually inactivate that gene See the non-uniform coloration of kernels in strains of Indian Corn

Mutation Rates Mutations occur spontaneously while DNA is being replicated, but fortunately special enzymes correct most of the mistakes. Each gene has a characteristic mutation rate Average rate for eukaryotes is between 10-4 and 10-6 per gene per generation Only mutations that arise in germ cells can be passed on to next generation

Mutagens Ionizing radiation (X rays) Nonionizing radiation (UV) Natural and synthetic chemicals - alkylating agents – add methyl or ethyl groups to DNA making it more vulnerable to mutations. (smoking)

Mutations If a mutation arises in a somatic cell, it will affect only that cell and will not be passed on to offspring. If, however, the mutation arises in a gamete, it may be passed on and thus enter the evolutionary arena. Either kind of mutation may prove to be harmful, beneficial, or neutral in its effects.