Genes to Proteins PDF

Genes to Proteins Genetic Information Flow in Prokaryotes (no nucleus): DNA mRNA transcription (RNA polymerase) protein translation (ribosomes) mRNA = messenger RNA ↳ polymers of amino acids ribosomes build proteins ribosomes build proteins • RNA polymerase: synthesizes RNA according to a...

Genes to Proteins Genetic Information Flow in Prokaryotes (no nucleus): DNA mRNA transcription (RNA polymerase) protein translation (ribosomes) mRNA = messenger RNA ↳ polymers of amino acids ribosomes build proteins ribosomes build proteins • RNA polymerase: synthesizes RNA according to a DNA template. • Transcription: DNA nucleotide-based language to an RNA nucleotide-based “language”. • Translation: RNA nucleotide-based language to an amino acid-based “language”. Genes to Proteins Genetic Information Flow in Eukaryotes (have a nucleus): ↳plants , animals etc , In the Nucleus: DNA pre-mRNA mRNA transcription processing (RNA polymerase) (splicing, modification) In the Cytosol: mRNA translation (ribosomes) protein ribosomes build proteins • RNA polymerase: synthesizes RNA according to a DNA template. • Transcription: DNA nucleotide-based language to an RNA nucleotide-based “language”. • Translation: RNA nucleotide-based language to an amino acid-based “language”. Genes to Proteins • • • • Product of transcription & translation is proteins. Proteins are polymers of amino acids. 20 protein amino acids. The information represented by DNA must somehow code for the protein 1° structure (= amino acid sequence). http://www.genome.gov/Pages/Hyperion//DIR/VIP/Glossary/Illustration/ami no_acid.shtml Genes to Proteins DNA Represents Information – the information is organized at various levels of structure: genome chromosomes decreasing scale genes codons nucleotides Codon – Three consecutive nucleotides that specify an amino acid (more details to come). Gene – One common definition of a gene is “a sequence of DNA/nucleotides/codons that specifies the primary structure of a protein (or a polypeptide)”; this is often called the “one gene-one protein” definition, or alternatively , the “one gene-one polypeptide” definition. rRNA and tRNA genes – The sections of DNA (nucleotides, codons) that specify the nucleotide sequences of rRNA and tRNA are also known as “genes”. So…….Another, More General, Definition of Gene – Sections of DNA that that code for a useful product, e.g. protein, tRNA, rRNA, or other forms of RNA. Genes to Proteins Structure of the Information • 20 protein amino acids must be encoded by DNA (via the genes). • There are only 4 DNA nucleotides (A, T, C, G);  can’t have a system where 1 nucleotide specifies 1 amino acid • How about two nucleotides to specify an amino acid?  4 x 4 = 16 possible combinations  still inadequate  cannot specify different 20 amino acids • Need a genetic code in which three nucleotides specify an amino acid:  4 x 4 x 4 = 64 possible combinations (> than the required 20)  the genetic code is a “triplet code” • Three nucleotides that specify an amino acid = a codon. • 64 codons. • But 64 is more than needed. • Genetic code is “redundant”;  18/20 protein amino acids are coded for by 2+ codons (exceptions are tryptophan and methionine) You do not need to know the exceptions. Genes to Proteins Table of the Genetic Code OpenStax Biology 2e Do not memorize this table • Usually shown as mRNA (notice all of the “U”s and lack of “T”s). • Three “stop codons” = stop translating here. • One “start codon” (AUG) that is also the codon for methionine;  the meaning of AUG is contextdependent. • Most codons specify amino acids. • The genetic code is very close to universal. This figure shows the genetic code for translating each nucleotide triplet in mRNA into an amino acid or a termination signal in a protein. AUG is the most common “start codon” and also codes for the amino acid methionine (the meaning of AUG is context-dependent). Genes to Proteins Nucleic Acids https://bio.libretexts.or g/@go/page/8394 Eukaryotic Transcription • In the nucleus. DNA DNA nucleotides (A, T, C, G) pre-mRNA RNA nucleotides (A, U, C, G) (5C sugars are different) • Enzyme: RNA polymerase; -assembles pre-mRNA according the instructions of the DNA -the DNA is the “template” • Nucleoplasm (sort of the cytosol of the nucleus) is kept stocked with RNA nucleotides. • RNA polymerase moves along the DNA strand;  adds one RNA nucleotide at a time • We that DNA is double-stranded, and that the strands are complementary;  if know the nucleotide sequence of one strand, you can work out the sequence of the other strand: DNA strand #1: GCCTATCCGAA A=T DNA strand #2: CGGATAGGCTT G≡C Genes to Proteins • One DNA strand acts as the template strand for RNA polymerase activity; DNA strand #1: GCCTATCCGAA pre-mRNA: CGGAUAGGCUU A=U G≡C The pre-mRNA is complementary to the DNA (same rules as for DNADNA, but U replaces T in RNA). OpenStax Biology 2e. Messenger RNA is a copy of protein-coding information in the coding strand of DNA, with the substitution of U in the RNA for T in the coding sequence. However, new RNA nucleotides base pair with the nucleotides of the template strand. RNA is synthesized in its 5'-3' direction, using the enzyme RNA polymerase. As the template is read, the DNA unwinds ahead of the polymerase and then rewinds behind it. Genes to Proteins Pre-mRNA Processing (Nucleus) DNA transcription (RNA polymerase) pre-mRNA mRNA processing (splicing, modification) • Pre-mRNA is also called the “primary transcript” (= the initial product of transcription in eukaryotes). • Pre-mRNA is produced by RNA polymerase. • Processing involves several steps: 1) Removal of introns – introns are parts of the gene that are not meant to be translated on the ribosome. 2) Exons must be spliced (glued) together – exons are the remaining parts of the gene that are meant to be translated on the ribosome. 3) Chemical modification of the ends of the transcript: addition of a 5’ (“five prime”) cap and a poly-A tail  prevents the mRNA from being degraded in the cytosol Genes to Proteins Pre-mRNA Processing (Nucleus) DNA transcription (RNA polymerase) pre-mRNA mRNA processing (splicing, modification) Introns are non-coding sequences; they must be removed. OpenStax Biology 2e. Primary transcript (= premRNA) is the product of RNA polymerase activity. Exons are coding sequences; they must be spliced (glued together). Only the exons remain after splicing. -S where info is Eukaryotic mRNA contains introns that must be spliced out. A 5' cap and 3' poly-A tail are also added. The 5’ cap contains modified “G”. Both the cap and the poly-A tail prevent the transcript from being broken down. Genes to Proteins Pre-mRNA Processing (Nucleus) DNA transcription (RNA polymerase) pre-mRNA mRNA processing (splicing, modification) OpenStax Biology 2e. Processing also includes addition of a “5’ cap”. The cap is composed of modified Gs. Processing also adds a “poly-A tail”. Processed transcript moves to the cytosol. Eukaryotic mRNA contains introns that must be spliced out. A 5' cap and 3' poly-A tail are also added. The 5’ cap contains modified “G”. Both the cap and the poly-A tail prevent the transcript from being broken down. Genes to Proteins Pre-mRNA Processing (Nucleus) – Another View UTR = untranslated region By Nastypatty - Own work, CC BY-SA 4.0, https://commons.wikimedia.org/w/index.php?curid=49282051 This region includes a nucleotide sequence recognized by the ribosome  leads to binding by the ribosome. Includes a “transcription terminator region”. You don’t need to know this. Digression – Two Nuclear Polymerases make come of DNA Two Nuclear Polymerases • DNA polymerase replicates DNA. • RNA polymerase produces RNA according to a DNA template (transcription). tan RNA Labe By Dhorspool at en.wikipedia, CC BY-SA 3.0, https://commons.wikimedia.org/w/index.php? curid=15183788 DNA Polymerase – replicates DNA template strand e.g. ATTGCTATTGCC replicated strand TAACGATAACGG A=T G≡C RNA Polymerase – produces RNA from a DNA template template strand e.g. ATTGCTATTGCC A=U RNA strand UAACGAUAACGG G≡C Genes to Proteins RNA is a Product of Transcription • There are several types of RNA; all are products of transcription. • We will consider three RNA types. • All RNAs are produced in the nucleus via RNA polymerase activity. • pre-mRNA  mRNA (= messenger RNA) – this is info (1° structure of a protein). • rRNA (= ribosomal RNA) – structural; component of ribosomes. • tRNA (= transfer RNA) – amino acid delivery system for translation. Ribosomes A ribosome is composed of two subunits: large and small. During translation, ribosomal subunits assemble together like a sandwich on the strand of mRNA. A long chain of linked amino acids emerges as the ribosome decodes the mRNA sequence into a polypeptide/protein. Genes to Proteins • • • • Ribosomes are protein-building machines. They can build any protein. The instructions for a particular protein are provided by the mRNA. Ribosomal subunits are assembled in the nucleus (nucleolus), from rRNA and specific ribosomal proteins. • Small and large subunits leave the nucleus separately  cytosol. • Need mRNA for ribosomal assembly  mRNA is sandwiched between the subunits. Functional Ribosome: small and large subunits, with mRNA ribosomal small subunit mRNA The ribosome moves along the mRNA. “Reads” the mRNA one codon at a time  adds the specified amino acid. ribosomal large subunit nascent polypeptide = polypeptide in progress Genes to Proteins Ribosomal subunits, with RNA in orange and yellow, and proteins in blue. https://pdb101.rcsb.org/motm/10 You are not expected to be able to draw this. Genes to Proteins Transfer RNA (tRNA) • tRNA is a product of transcription (tRNA genes). • No protein. • Many types of tRNA. • Has an approximate cloverleaf-like pattern in two dimensions. • Has two important parts: 1) anti-codon 2) amino acid binding site Amino acid binding site H bonds between RNA nucleotides You do not need to be able to draw this in detail. Modified from Berg et al. (2019) RNA Biology 16: 1-12 Genes to Proteins Three dimensional view of tRNA. You do not need to be able to draw this. This is a space-filling model of a tRNA molecule that adds the amino acid phenylalanine to a growing polypeptide chain. The anticodon AAG binds the Codon UUC on the mRNA. The amino acid phenylalanine is attached to the other end of the tRNA. anticodon AAG is complementary to the mRNA codon UUC = phenylalanine OpenStax Biology 2e Genes to Proteins Transfer RNA (tRNA) • Amino acid binding site exhibits specificity;  binds one of the 20 protein amino acids • Anticodon is complementary to an mRNA codon. • Let’s assume that anticodon is AAG. • AAG is complementary to UUC. • UUC is an mRNA codon for phenyalanine. • Have a matching system: if the anticodon and the codon are complementary, then the ribosome adds the amino acid in the amino acid binding site to the growing polypeptide. • If the anticodon is AAG, then the binding site is for phenyalanine. Amino acid binding site Modified from Berg et al. (2019) RNA Biology 16: 1-12 Genes to Proteins • tRNAs with bound amino acids are called aminoacyl tRNA or charged tRNA. • Cytosolic amino acid pool (cytosol is kept stocked with amino acids). • tRNAs can be used repeatedly. Alcamo (2001) Fundamentals of Microbiology 6th ed. Jones and Bartlett. transcription (RNA polymerase) cytosolic amino acid pool nucleus cytosol aminoacyl tRNA (= tRNA with bound amino acid) tRNA translation (occurring on ribosome) tRNA pre-mRNA mRNA pre-mRNA splicing/processing mRNA ssu protein rRNA protein ssu polypeptide being built (in progress) lsu lsu strand of DNA (complementary strand not shown) nuclear envelope (pores not shown) ssu = ribosomal small subunit lsu = ribosomal large subunit Summary Diagram for Eukaryote Genetic Information Flow • Does not show the ribosomes, tRNA or cytosolic amino acid pool. https://www.nature.com/scitable/topicpage/geneexpression-14121669/ An overview of the flow of information from DNA to protein in a eukaryote. First, both coding and noncoding regions of DNA are transcribed into mRNA. Some regions are removed (introns) during initial mRNA processing. The remaining exons are then spliced together, and the spliced mRNA molecule (red) is prepared for export out of the nucleus through addition of an endcap (sphere) and a polyA tail. Once in the cytoplasm, the mRNA can be used to construct a protein. Genes to Proteins Cellular Energy Costs of Protein Synthesis are Large • There is a cytosolic enzyme that adds the amino acids to the amino acid binding site on the tRNA;  with an ATP cost = 2 ATP per amino acid • Ribosomes also require energy;  several ATP equivalents per amino acid added (the ribosome uses GTP). • Transcription also has an ATP cost;  2 ATP per RNA nucleotide added to mRNA by RNA polymerase • Estimated that protein synthesis accounts for ~20% of basal metabolic rate: 1) adding amino acids to tRNA  aminoacyl tRNA 2) adding amino acids to the growing polypeptide (ribosome) 3) production of RNA by transcription (RNA polymerase in the nucleus) Genes to Proteins – Regulation of Transcription • “Gene expression” is the formation of a gene product from a gene; -polypeptide/protein -RNA (rRNA, tRNA, and other RNAs that we have not discussed) • Gene expression is tightly-regulated. • Different genes have different expression patterns in in different tissues. • Transcriptional regulation is an important part of regulating gene expression. • Transcription factors are proteins that regulate transcription; -they are themselves products of gene expression -human genome estimated to contain ~1600 genes for transcription factors (out ~21,000 genes) • Transcription factors bind to promoters of genes, and regulate the activity of RNA polymerase. • Promotors are part of the non-transcribed portion of a gene (this different from an untranslated region); -promotors have a transcription regulatory function, and are affected by the binding of transcription factors Genes to Proteins – Regulation of Transcription • Transcription factors may promote transcription by RNA polymerase (positive transcription factor) or inhibit transcription (negative transcription factor). Simplified view of promoters and transcription factors: transcription factor RNA polymerase (responsible for transcription) transcription leads to pre-mRNA pre-mRNA (product of transcription) promoter (non-transcribed) transcribed portion of gene, used as the template for pre-mRNA gene The promoter is a non-transcribed portion of the gene; it has a regulatory function (regulates transcription by RNA polymerase). Transcription factors (which are proteins) bind to specific regions of the promoter, and may promote or inhibit transcription. Genes to Proteins – Regulation of Transcription • The TATA box (TATAAA) is a common nucleotide sequence in promoters where transcription factors tend to bind. • There may be multiple (many) transcription factors that simultaneously bind to a single promotor. • The binding and activity of RNA polymerase is regulated by the transcription factors. A generalized promoter of a gene transcribed by RNA polymerase II is shown. Transcription factors recognize the promoter. RNA polymerase II then binds and forms the transcription initiation complex. You will not be asked to be able to draw this, or memorize the various transcription factors. OpenStax Biology 2e Genes to Proteins – Regulation of Transcription • The TATA box (TATAAA) is a common nucleotide sequence in promoters where transcription factors tend to bind. • There may be multiple (many) transcription factors that simultaneously bind to a single promotor. • The binding and activity of RNA polymerase is regulated by the transcription factors. A generalized promoter of a gene transcribed by RNA polymerase II is shown. Transcription factors recognize the promoter. RNA polymerase II then binds and forms the transcription initiation complex. You will not be asked to be able to draw this, or memorize the various transcription factors. OpenStax Biology 2e