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Molecular genetics.ppt

The document discusses the flow of genetic information from DNA to protein. It describes how DNA contains the genetic code in the form of nucleotide sequences. During transcription, DNA directs the synthesis of messenger RNA (mRNA). Translation then uses the mRNA to synthesize polypeptides through the ribosome. George Beadle and Edward Tatum provided evidence supporting the "one gene-one enzyme" hypothesis through experiments with bread mold mutants. Their work showed that each gene directs the production of a specific enzyme.

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Molecular genetics From Gene to Protein PowerPoint Lectures for Biology, Seventh Edition Neil Campbell and Jane Reece Lectures by Chris Romero Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  Overview: The Flow of Genetic Information •  The information content of DNA –  Is in the form of specific sequences of nucleotides along the DNA strands Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  The DNA inherited by an organism –  Leads to specific traits by dictating the synthesis of proteins •  The process by which DNA directs protein synthesis, gene expression –  Includes two stages, called transcription and translation Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  The ribosome –  Is part of the cellular machinery for translation, polypeptide synthesis Figure 17.1 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  Concept 17.1: Genes specify proteins via transcription and translation Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Evidence from the Study of Metabolic Defects •  In 1909, British physician Archibald Garrod –  Was the first to suggest that genes dictate phenotypes through enzymes that catalyze specific chemical reactions in the cell Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Nutritional Mutants in Neurospora: Scientific Inquiry •  Beadle and Tatum causes bread mold to mutate with X-rays –  Creating mutants that could not survive on minimal medium Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  Using genetic crosses –  They determined that their mutants fell into three classes, each mutated in a different gene EXPERIMENT RESULTS Working with the mold Neurospora crassa, George Beadle and Edward Tatum had isolated mutants requiring arginine in their growth medium and had shown genetically that these mutants fell into three classes, each defective in a different gene. From other considerations, they suspected that the metabolic pathway of arginine biosynthesis included the precursors ornithine and citrulline. Their most famous experiment, shown here, tested both their one gene–one enzyme hypothesis and their postulated arginine pathway. In this experiment, they grew their three classes of mutants under the four different conditions shown in the Results section below. The wild-type strain required only the minimal medium for growth. The three classes of mutants had different growth requirements Wild type Minimal medium (MM) (control) MM + Ornithine MM + Citrulline Figure 17.2 MM + Arginine (control) Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Class I Mutants Class II Mutants Class III Mutants
CONCLUSION From the growth patterns of the mutants, Beadle and Tatum deduced that each mutant was unable to carry out one step in the pathway for synthesizing arginine, presumably because it lacked the necessary enzyme. Because each of their mutants was mutated in a single gene, they concluded that each mutated gene must normally dictate the production of one enzyme. Their results supported the one gene–one enzyme hypothesis and also confirmed the arginine pathway. (Notice that a mutant can grow only if supplied with a compound made after the defective step.) Wild type Precursor Gene A Class I Mutants (mutation in gene A) Precursor Precursor A A Ornithine Ornithine Ornithine B B B Citrulline Citrulline Citrulline C C C Arginine Arginine Arginine Enzyme A Ornithine Gene B Enzyme B Citrulline Gene C Enzyme C Arginine Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Class II Mutants (mutation in gene B) Class III Mutants (mutation in gene C) Precursor A
•  Beadle and Tatum developed the “one gene– one enzyme hypothesis” –  Which states that the function of a gene is to dictate the production of a specific enzyme Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
The Products of Gene Expression: A Developing Story •  As researchers learned more about proteins –  The made minor revision to the one gene–one enzyme hypothesis •  Genes code for polypeptide chains or for RNA molecules Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Basic Principles of Transcription and Translation •  Transcription –  Is the synthesis of RNA under the direction of DNA –  Produces messenger RNA (mRNA) •  Translation –  Is the actual synthesis of a polypeptide, which occurs under the direction of mRNA –  Occurs on ribosomes Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  In prokaryotes –  Transcription and translation occur together TRANSCRIPTION DNA mRNA Ribosome TRANSLATION Polypeptide (a) Prokaryotic cell. In a cell lacking a nucleus, mRNA produced by transcription is immediately translated without additional processing. Figure 17.3a Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  In eukaryotes –  RNA transcripts are modified before becoming true mRNA Nuclear envelope DNA TRANSCRIPTION Pre-mRNA RNA PROCESSING mRNA Ribosome TRANSLATION Polypeptide Figure 17.3b Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings (b) Eukaryotic cell. The nucleus provides a separate compartment for transcription. The original RNA transcript, called pre-mRNA, is processed in various ways before leaving the nucleus as mRNA.
•  Cells are governed by a cellular chain of command –  DNA → RNA → protein Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
The Genetic Code •  How many bases correspond to an amino acid? Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Codons: Triplets of Bases •  Genetic information –  Is encoded as a sequence of nonoverlapping base triplets, or codons Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  During transcription –  The gene determines the sequence of bases along the length of an mRNA molecule Gene 2 DNA molecule Gene 1 Gene 3 DNA strand 3ʹ′ 5ʹ′ A C C A A A C C G A G T (template) TRANSCRIPTION mRNA 5ʹ′ U G G U U U G G C U C A Codon TRANSLATION Protein Figure 17.4 Trp Amino acid Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Phe Gly Ser 3ʹ′
Cracking the Code •  A codon in messenger RNA Figure 17.5 Second mRNA base U C A UAU UUU UCU Tyr Phe UAC UUC UCC U UUA UCA Ser UAA Stop UAG Stop UUG Leu UCG CUU CUC C CUA CUG CCU CCC Leu CCA CCG Pro AUU AUC A AUA AUG ACU ACC ACA ACG Thr GUU G GUC GUA GUG lle Met or start GCU GCC Val GCA GCG Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Ala G U UGU Cys UGC C UGA Stop A UGG Trp G U CAU CGU His CAC CGC C Arg CAA CGA A Gln CAG CGG G U AAU AGU Asn AAC AGC Ser C A AAA AGA Lys AAG AGG Arg G U GAU GGU C GAC Asp GGC Gly GAA GGA A Glu GGG GAG G Third mRNA base (3ʹ′ end) First mRNA base (5ʹ′ end) –  Is either translated into an amino acid or serves as a translational stop signal
•  Codons must be read in the correct reading frame –  For the specified polypeptide to be produced Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Evolution of the Genetic Code •  The genetic code is nearly universal –  Shared by organisms from the simplest bacteria to the most complex animals Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  In laboratory experiments –  Genes can be transcribed and translated after being transplanted from one species to another Figure 17.6 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  Concept 17.2: Transcription is the DNAdirected synthesis of RNA: a closer look Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Molecular Components of Transcription •  RNA synthesis –  Is catalyzed by RNA polymerase, which pries the DNA strands apart and hooks together the RNA nucleotides –  Follows the same base-pairing rules as DNA, except that in RNA, uracil substitutes for thymine Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Synthesis of an RNA Transcript •  The stages of transcription are Promoter –  Initiation Transcription unit 5ʹ′ 3ʹ′ Start point RNA polymerase –  Elongation –  Termination 3ʹ′ 5ʹ′ DNA 1 Initiation. After RNA polymerase binds to the promoter, the DNA strands unwind, and the polymerase initiates RNA synthesis at the start point on the template strand. 5ʹ′ 3ʹ′ Unwound DNA 3ʹ′ 5ʹ′ Template strand of DNA transcript 2 Elongation. The polymerase moves downstream, unwinding the DNA and elongating the RNA transcript 5ʹ′ → 3 ʹ′. In the wake of transcription, the DNA strands re-form a double helix. Rewound RNA RNA 5ʹ′ 3ʹ′ 3ʹ′ 5ʹ′ 3ʹ′ 5ʹ′ RNA transcript 3 Termination. Eventually, the RNA transcript is released, and the polymerase detaches from the DNA. 5ʹ′ 3ʹ′ 3ʹ′ 5ʹ′ 5ʹ′ Figure 17.7 Completed RNA transcript Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 3ʹ′
Non-template strand of DNA Elongation RNA nucleotides RNA polymerase A 3ʹ′ T C C C A A T U 3ʹ′ end U 5ʹ′ C A T E A G G C A G T A Newly made RNA Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings C T Direction of transcription (“downstream”) 5ʹ′ G A Template strand of DNA
RNA Polymerase Binding and Initiation of Transcription •  Promoters signal the initiation of RNA synthesis •  Transcription factors –  Help eukaryotic RNA polymerase recognize promoter sequences TRANSCRIPTION RNA PROCESSING 1 Eukaryotic promoters DNA Pre-mRNA mRNA TRANSLATION Ribosome Polypeptide 5ʹ′ 3ʹ′ Promoter 3ʹ′ 5ʹ′ T A T A A AA AT A T T T T TATA box Start point Template DNA strand Several transcription factors 2 Transcription factors 5ʹ′ 3ʹ′ 3ʹ′ 5ʹ′ 3 Additional transcription factors RNA polymerase II 5ʹ′ 3ʹ′ Figure 17.8 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Transcription factors 3ʹ′ 5ʹ′ 5ʹ′ RNA transcript Transcription initiation complex
Elongation of the RNA Strand •  As RNA polymerase moves along the DNA –  It continues to untwist the double helix, exposing about 10 to 20 DNA bases at a time for pairing with RNA nucleotides Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Termination of Transcription •  The mechanisms of termination –  Are different in prokaryotes and eukaryotes Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  Concept 17.3: Eukaryotic cells modify RNA after transcription •  Enzymes in the eukaryotic nucleus –  Modify pre-mRNA in specific ways before the genetic messages are dispatched to the cytoplasm Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Alteration of mRNA Ends •  Each end of a pre-mRNA molecule is modified in a particular way –  The 5ʹ′ end receives a modified nucleotide cap –  The 3ʹ′ end gets a poly-A tail A modified guanine nucleotide added to the 5ʹ′ end TRANSCRIPTION RNA PROCESSING 50 to 250 adenine nucleotides added to the 3ʹ′ end DNA Pre-mRNA mRNA 5ʹ′ Protein-coding segment G P P P AAUAAA Ribosome TRANSLATION 5ʹ′ Cap Polypeptide Polyadenylation signal 5ʹ′ UTR Start codon Stop codon Figure 17.9 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 3ʹ′ UTR 3ʹ′ AAA…AAA Poly-A tail
Split Genes and RNA Splicing •  RNA splicing –  Removes introns and joins exons TRANSCRIPTION RNA PROCESSING DNA Pre-mRNA 5ʹ′ Exon Intron Pre-mRNA 5ʹ′ Cap 30 31 1 Coding segment mRNA Ribosome Intron Exon Exon 3ʹ′ Poly-A tail 104 105 146 Introns cut out and exons spliced together TRANSLATION Polypeptide mRNA 5ʹ′ Cap 1 3ʹ′ UTR Figure 17.10 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Poly-A tail 146 3ʹ′ UTR
•  Is carried out by spliceosomes in some cases 5ʹ′ 1 RNA transcript (pre-mRNA) Intron Exon 1 Exon 2 Protein Other proteins snRNA snRNPs Spliceosome 2 5ʹ′ Spliceosome components 3 Figure 17.11 5ʹ′ mRNA Exon 1 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Exon 2 Cut-out intron
Ribozymes •  Ribozymes –  Are catalytic RNA molecules that function as enzymes and can splice RNA Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
The Functional and Evolutionary Importance of Introns •  The presence of introns –  Allows for alternative RNA splicing Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  Proteins often have a modular architecture –  Consisting of discrete structural and functional regions called domains •  In many cases –  Different exons code for the different domains in a protein Gene DNA Exon 1 Intron Exon 2 Transcription RNA processing Intron Exon 3 Translation Domain 3 Domain 2 Domain 1 Figure 17.12 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Polypeptide
•  Concept 17.4: Translation is the RNA-directed synthesis of a polypeptide: a closer look Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Molecular Components of Translation •  A cell translates an mRNA message into protein –  With the help of transfer RNA (tRNA) Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  Translation: the basic concept TRANSCRIPTION DNA mRNA Ribosome TRANSLATION Polypeptide Amino acids Polypeptide tRNA with amino acid Ribosome attached Ph e Gly tRNA A GC A A A U G G U U U G G C Codons 5ʹ′ Figure 17.13 mRNA Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Anticodon 3ʹ′
•  Molecules of tRNA are not all identical –  Each carries a specific amino acid on one end –  Each has an anticodon on the other end Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
The Structure and Function of Transfer RNA •  A tRNA molecule A –  Consists of a single RNA strand C that is only C about 80 nucleotides long –  Is roughly L-shaped 3ʹ′ A C C A 5ʹ′ C G G C C G U G U A A U A U U C UA C A C AG * G * G U G U * C C * * U C * * G AG C (a) Two-dimensional structure. The four base-paired regions and three G C U A loops are characteristic of all tRNAs, as is the base sequence of the * G amino acid attachment site at the 3ʹ′ end. The anticodon triplet is A A* unique to each tRNA type. (The asterisks mark bases that have been C U * chemically modified, a characteristic of tRNA.) A G A Figure 17.14a Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Amino acid attachment site Anticodon C U C G A G A G * * G A G G Hydrogen bonds
5ʹ′ 3ʹ′ Amino acid attachment site Hydrogen bonds A AG 3ʹ′ Anticodon (b) Three-dimensional structure Figure 17.14b Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 5ʹ′ Anticodon (c) Symbol used in this book
•  A specific enzyme called an aminoacyl-tRNA synthetase –  Joins each amino acid to the correct tRNA Amino acid Aminoacyl-tRNA synthetase (enzyme) 1 Active site binds the amino acid and ATP. P P P Adenosine ATP 2 ATP loses two P groups and joins amino acid as AMP. P Pyrophosphate Pi Phosphates 3 Appropriate tRNA covalently Bonds to amino Acid, displacing AMP. P Adenosine Pi Pi tRNA P Adenosine AMP 4 Activated amino acid is released by the enzyme. Figure 17.15 Aminoacyl tRNA (an “activated amino acid”) Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Ribosomes •  Ribosomes –  Facilitate the specific coupling of tRNA anticodons with mRNA codons during protein synthesis Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  The ribosomal subunits –  Are constructed of proteins and RNA molecules named ribosomal RNA or rRNA DNA TRANSCRIPTION mRNA Ribosome TRANSLATION Polypeptide Growing polypeptide Exit tunnel tRNA molecules Large subunit E P A Small subunit 5ʹ′ mRNA Figure 17.16a 3ʹ′ (a) Computer model of functioning ribosome. This is a model of a bacterial ribosome, showing its overall shape. The eukaryotic ribosome is roughly similar. A ribosomal subunit is an aggregate of ribosomal RNA molecules and proteins. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  The ribosome has three binding sites for tRNA –  The P site –  The A site –  The E site P site (Peptidyl-tRNA binding site) A site (AminoacyltRNA binding site) E site (Exit site) Large subunit E mRNA binding site Figure 17.16b P A Small subunit (b) Schematic model showing binding sites. A ribosome has an mRNA binding site and three tRNA binding sites, known as the A, P, and E sites. This schematic ribosome will appear in later diagrams. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Amino end Growing polypeptide Next amino acid to be added to polypeptide chain tRNA 3ʹ′ mRNA 5ʹ′ Codons (c) Schematic model with mRNA and tRNA. A tRNA fits into a binding site when its anticodon base-pairs with an mRNA codon. The P site holds the tRNA attached to the growing polypeptide. The A site holds the tRNA carrying the next amino acid to be added to the polypeptide chain. Discharged tRNA leaves via the E site. Figure 17.16c Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Building a Polypeptide •  We can divide translation into three stages –  Initiation –  Elongation –  Termination Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Ribosome Association and Initiation of Translation •  The initiation stage of translation –  Brings together mRNA, tRNA bearing the first amino acid of the polypeptide, and two subunits of a ribosome P site 3ʹ′ U A C 5ʹ′ 5ʹ′ A U G 3ʹ′ Initiator tRNA GTP GDP E mRNA 5ʹ′ Start codon mRNA binding site Figure 17.17 3ʹ′ Small ribosomal subunit 1 A small ribosomal subunit binds to a molecule of mRNA. In a prokaryotic cell, the mRNA binding site on this subunit recognizes a specific nucleotide sequence on the mRNA just upstream of the start codon. An initiator tRNA, with the anticodon UAC, base-pairs with the start codon, AUG. This tRNA carries the amino acid methionine (Met). Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Large ribosomal subunit 5ʹ′ A 3ʹ′ Translation initiation complex 2 The arrival of a large ribosomal subunit completes the initiation complex. Proteins called initiation factors (not shown) are required to bring all the translation components together. GTP provides the energy for the assembly. The initiator tRNA is in the P site; the A site is available to the tRNA bearing the next amino acid.
Elongation of the Polypeptide Chain •  In the elongation stage of translation –  Amino acids are added one by one to the preceding amino acid TRANSCRIPTION Amino end of polypeptide DNA mRNA Ribosome TRANSLATION Polypeptide mRNA Ribosome ready for next aminoacyl tRNA E 3ʹ′ P A site site 5ʹ′ 1 Codon recognition. The anticodon of an incoming aminoacyl tRNA base-pairs with the complementary mRNA codon in the A site. Hydrolysis of GTP increases the accuracy and efficiency of this step. 2 GTP 2 GDP E E P Figure 17.18 3 Translocation. The ribosome translocates the tRNA in the A site to the P site. The empty tRNA in the P site is moved to the E site, where it is released. The mRNA moves along with its bound tRNAs, bringing the next codon to be translated into the A site. P A GDP GTP Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings E P A A 2 Peptide bond formation. An rRNA molecule of the large subunit catalyzes the formation of a peptide bond between the new amino acid in the A site and the carboxyl end of the growing polypeptide in the P site. This step attaches the polypeptide to the tRNA in the A site.
Termination of Translation •  The final stage of translation is termination –  When the ribosome reaches a stop codon in the mRNA Release factor Free polypeptide 5ʹ′ 3ʹ′ 3ʹ′ 5ʹ′ 5ʹ′ 3ʹ′ Stop codon (UAG, UAA, or UGA) 1 When a ribosome reaches a stop 2 The release factor hydrolyzes 3 The two ribosomal subunits codon on mRNA, the A site of the the bond between the tRNA in and the other components of ribosome accepts a protein called the P site and the last amino the assembly dissociate. a release factor instead of tRNA. acid of the polypeptide chain. The polypeptide is thus freed from the ribosome. Figure 17.19 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Polyribosomes •  A number of ribosomes can translate a single mRNA molecule simultaneously –  Forming a polyribosome Completed polypeptide Growing polypeptides Incoming ribosomal subunits Start of mRNA (5ʹ′ end) Polyribosom End of mRNA (3ʹ′ end) (a) An mRNA molecule is generally translated simultaneously by several ribosomes in clusters called polyribosomes. e Ribosomes mRNA 0.1 µm Figure 17.20a, b (b) This micrograph shows a large polyribosome in a prokaryotic cell (TEM). Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Completing and Targeting the Functional Protein •  Polypeptide chains –  Undergo modifications after the translation process Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Protein Folding and Post-Translational Modifications •  After translation –  Proteins may be modified in ways that affect their three-dimensional shape Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Targeting Polypeptides to Specific Locations •  Two populations of ribosomes are evident in cells –  Free and bound •  Free ribosomes in the cytosol –  Initiate the synthesis of all proteins Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  Proteins destined for the endomembrane system or for secretion –  Must be transported into the ER –  Have signal peptides to which a signalrecognition particle (SRP) binds, enabling the translation ribosome to bind to the ER Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  The signal mechanism for targeting proteins to the ER 1 Polypeptide synthesis begins on a free ribosome in the cytosol. 2 An SRP binds to the signal peptide, halting synthesis momentarily. 3 The SRP binds to a receptor protein in the ER membrane. This receptor is part of a protein complex (a translocation complex) that has a membrane pore and a signal-cleaving enzyme. 4 The SRP leaves, and 5 The signalthe polypeptide resumes cleaving growing, meanwhile enzyme translocating across the cuts off the membrane. (The signal signal peptide. peptide stays attached to the membrane.) 6 The rest of the completed polypeptide leaves the ribosome and folds into its final conformation. Ribosome mRNA Signal peptide Signalrecognition particle (SRP) SRP receptor CYTOSOL protein ERLUMEN Translocation complex Figure 17.21 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Signal peptide removed ER membrane Protein
•  Concept 17.5: RNA plays multiple roles in the cell: a review •  RNA –  Can hydrogen-bond to other nucleic acid molecules –  Can assume a specific three-dimensional shape –  Has functional groups that allow it to act as a catalyst Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  Types of RNA in a Eukaryotic Cell Table 17.1 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  Concept 17.6: Comparing gene expression in prokaryotes and eukaryotes reveals key differences •  Prokaryotic cells lack a nuclear envelope –  Allowing translation to begin while transcription is still in progress RNA polymerase DNA mRNA Polyribosome RNA polymerase Direction of transcription DNA Polyribosome Polypeptide (amino end) Ribosome Figure 17.22 0.25 µm mRNA (5ʹ′ end) Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  In a eukaryotic cell –  The nuclear envelope separates transcription from translation –  Extensive RNA processing occurs in the nucleus Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  Concept 17.7: Point mutations can affect protein structure and function •  Mutations –  Are changes in the genetic material of a cell •  Point mutations –  Are changes in just one base pair of a gene Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  The change of a single nucleotide in the DNA’s template strand –  Leads to the production of an abnormal protein Wild-type hemoglobin DNA 3ʹ′ Mutant hemoglobin DNA C T T G U A 5ʹ′ In the DNA, the mutant template strand has an A where the wild-type template has a T. The mutant mRNA has a U instead of an A in one codon. 3ʹ′ 5ʹ′ T C A mRNA mRNA G A A 5ʹ′ 3ʹ′ 5ʹ′ 3ʹ′ Normal hemoglobin Sickle-cell hemoglobin Glu Val Figure 17.23 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings The mutant (sickle-cell) hemoglobin has a valine (Val) instead of a glutamic acid (Glu).
Types of Point Mutations •  Point mutations within a gene can be divided into two general categories –  Base-pair substitutions –  Base-pair insertions or deletions Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Substitutions •  A base-pair substitution –  Is the replacement of one nucleotide and its partner with another pair of nucleotides –  Can cause missense or nonsense Wild type mRNA Protein 5ʹ′ A U G Met A A G U U U GG C U A A Lys Phe Gly 3ʹ′ Stop Amino end Carboxyl end Base-pair substitution No effect on amino acid sequence U instead of C A U G A A G U U U G G U U A A Met Lys Missense Phe Gly Stop A instead of G A U G A A G U U U A G U U A A Met Lys Phe Ser Stop Nonsense U instead of A A U G U A G U U U G G C U A A Figure 17.24 Met Stop Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
Insertions and Deletions •  Insertions and deletions –  Are additions or losses of nucleotide pairs in a gene –  May produce frameshift mutations Wild type mRNA Protein 5ʹ′ A U GA A GU U U GG C U A A Met Lys Gly Phe Stop Amino end Carboxyl end Base-pair insertion or deletion Frameshift causing immediate nonsense Extra U AU GU A AG U U U G GC U A Met Stop Frameshift causing extensive missense U Missing A U G A A GU U G G C U A A Met Lys Leu Ala Insertion or deletion of 3 nucleotides: no frameshift but extra or missing amino acid A A G Missing A U G U U U G G C U A A Figure 17.25 Met Phe Gly Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Stop 3ʹ′
Mutagens •  Spontaneous mutations –  Can occur during DNA replication, recombination, or repair Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  Mutagens –  Are physical or chemical agents that can cause mutations Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
What is a gene? revisiting the question •  A gene –  Is a region of DNA whose final product is either a polypeptide or an RNA molecule Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
•  A summary of transcription and translation in a eukaryotic cell DNA TRANSCRIPTION 1 RNA is transcribed from a DNA template. 3ʹ′ RNA transcript 5ʹ′ RNA polymerase RNA PROCESSING Exon 2 In eukaryotes, the RNA transcript (premRNA) is spliced and modified to produce mRNA, which moves from the nucleus to the cytoplasm. RNA transcript (pre-mRNA) Intron Aminoacyl-tRNA synthetase NUCLEUS Amino acid tRNA FORMATION OF INITIATION COMPLEX CYTOPLASM 3 After leaving the nucleus, mRNA attaches to the ribosome. mRNA AMINO ACID ACTIVATION 4 Each amino acid attaches to its proper tRNA with the help of a specific enzyme and ATP. Growing polypeptide Activated amino acid Ribosomal subunits 5ʹ′ TRANSLATION A succession of tRNAs add their amino acids to the polypeptide chain Anticodon as the mRNA is moved through the ribosome one codon at a time. (When completed, the polypeptide is released from the ribosome.) 5 E A AAA UG GU U U A U G Codon Figure 17.26 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Ribosome

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Molecular genetics.ppt

  • 1.
    Molecular genetics From Geneto Protein PowerPoint Lectures for Biology, Seventh Edition Neil Campbell and Jane Reece Lectures by Chris Romero Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 2.
    •  Overview: TheFlow of Genetic Information •  The information content of DNA –  Is in the form of specific sequences of nucleotides along the DNA strands Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 3.
    •  The DNAinherited by an organism –  Leads to specific traits by dictating the synthesis of proteins •  The process by which DNA directs protein synthesis, gene expression –  Includes two stages, called transcription and translation Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 4.
    •  The ribosome – Is part of the cellular machinery for translation, polypeptide synthesis Figure 17.1 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 5.
    •  Concept 17.1:Genes specify proteins via transcription and translation Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 6.
    Evidence from theStudy of Metabolic Defects •  In 1909, British physician Archibald Garrod –  Was the first to suggest that genes dictate phenotypes through enzymes that catalyze specific chemical reactions in the cell Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 7.
    Nutritional Mutants inNeurospora: Scientific Inquiry •  Beadle and Tatum causes bread mold to mutate with X-rays –  Creating mutants that could not survive on minimal medium Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 8.
    •  Using geneticcrosses –  They determined that their mutants fell into three classes, each mutated in a different gene EXPERIMENT RESULTS Working with the mold Neurospora crassa, George Beadle and Edward Tatum had isolated mutants requiring arginine in their growth medium and had shown genetically that these mutants fell into three classes, each defective in a different gene. From other considerations, they suspected that the metabolic pathway of arginine biosynthesis included the precursors ornithine and citrulline. Their most famous experiment, shown here, tested both their one gene–one enzyme hypothesis and their postulated arginine pathway. In this experiment, they grew their three classes of mutants under the four different conditions shown in the Results section below. The wild-type strain required only the minimal medium for growth. The three classes of mutants had different growth requirements Wild type Minimal medium (MM) (control) MM + Ornithine MM + Citrulline Figure 17.2 MM + Arginine (control) Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Class I Mutants Class II Mutants Class III Mutants
  • 9.
    CONCLUSION From the growthpatterns of the mutants, Beadle and Tatum deduced that each mutant was unable to carry out one step in the pathway for synthesizing arginine, presumably because it lacked the necessary enzyme. Because each of their mutants was mutated in a single gene, they concluded that each mutated gene must normally dictate the production of one enzyme. Their results supported the one gene–one enzyme hypothesis and also confirmed the arginine pathway. (Notice that a mutant can grow only if supplied with a compound made after the defective step.) Wild type Precursor Gene A Class I Mutants (mutation in gene A) Precursor Precursor A A Ornithine Ornithine Ornithine B B B Citrulline Citrulline Citrulline C C C Arginine Arginine Arginine Enzyme A Ornithine Gene B Enzyme B Citrulline Gene C Enzyme C Arginine Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Class II Mutants (mutation in gene B) Class III Mutants (mutation in gene C) Precursor A
  • 10.
    •  Beadle andTatum developed the “one gene– one enzyme hypothesis” –  Which states that the function of a gene is to dictate the production of a specific enzyme Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 11.
    The Products ofGene Expression: A Developing Story •  As researchers learned more about proteins –  The made minor revision to the one gene–one enzyme hypothesis •  Genes code for polypeptide chains or for RNA molecules Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 12.
    Basic Principles ofTranscription and Translation •  Transcription –  Is the synthesis of RNA under the direction of DNA –  Produces messenger RNA (mRNA) •  Translation –  Is the actual synthesis of a polypeptide, which occurs under the direction of mRNA –  Occurs on ribosomes Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 13.
    •  In prokaryotes – Transcription and translation occur together TRANSCRIPTION DNA mRNA Ribosome TRANSLATION Polypeptide (a) Prokaryotic cell. In a cell lacking a nucleus, mRNA produced by transcription is immediately translated without additional processing. Figure 17.3a Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 14.
    •  In eukaryotes – RNA transcripts are modified before becoming true mRNA Nuclear envelope DNA TRANSCRIPTION Pre-mRNA RNA PROCESSING mRNA Ribosome TRANSLATION Polypeptide Figure 17.3b Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings (b) Eukaryotic cell. The nucleus provides a separate compartment for transcription. The original RNA transcript, called pre-mRNA, is processed in various ways before leaving the nucleus as mRNA.
  • 15.
    •  Cells aregoverned by a cellular chain of command –  DNA → RNA → protein Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 16.
    The Genetic Code • How many bases correspond to an amino acid? Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 17.
    Codons: Triplets ofBases •  Genetic information –  Is encoded as a sequence of nonoverlapping base triplets, or codons Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 18.
    •  During transcription – The gene determines the sequence of bases along the length of an mRNA molecule Gene 2 DNA molecule Gene 1 Gene 3 DNA strand 3ʹ′ 5ʹ′ A C C A A A C C G A G T (template) TRANSCRIPTION mRNA 5ʹ′ U G G U U U G G C U C A Codon TRANSLATION Protein Figure 17.4 Trp Amino acid Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Phe Gly Ser 3ʹ′
  • 19.
    Cracking the Code • A codon in messenger RNA Figure 17.5 Second mRNA base U C A UAU UUU UCU Tyr Phe UAC UUC UCC U UUA UCA Ser UAA Stop UAG Stop UUG Leu UCG CUU CUC C CUA CUG CCU CCC Leu CCA CCG Pro AUU AUC A AUA AUG ACU ACC ACA ACG Thr GUU G GUC GUA GUG lle Met or start GCU GCC Val GCA GCG Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Ala G U UGU Cys UGC C UGA Stop A UGG Trp G U CAU CGU His CAC CGC C Arg CAA CGA A Gln CAG CGG G U AAU AGU Asn AAC AGC Ser C A AAA AGA Lys AAG AGG Arg G U GAU GGU C GAC Asp GGC Gly GAA GGA A Glu GGG GAG G Third mRNA base (3ʹ′ end) First mRNA base (5ʹ′ end) –  Is either translated into an amino acid or serves as a translational stop signal
  • 20.
    •  Codons mustbe read in the correct reading frame –  For the specified polypeptide to be produced Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 21.
    Evolution of theGenetic Code •  The genetic code is nearly universal –  Shared by organisms from the simplest bacteria to the most complex animals Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 22.
    •  In laboratoryexperiments –  Genes can be transcribed and translated after being transplanted from one species to another Figure 17.6 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 23.
    •  Concept 17.2:Transcription is the DNAdirected synthesis of RNA: a closer look Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 24.
    Molecular Components ofTranscription •  RNA synthesis –  Is catalyzed by RNA polymerase, which pries the DNA strands apart and hooks together the RNA nucleotides –  Follows the same base-pairing rules as DNA, except that in RNA, uracil substitutes for thymine Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 25.
    Synthesis of anRNA Transcript •  The stages of transcription are Promoter –  Initiation Transcription unit 5ʹ′ 3ʹ′ Start point RNA polymerase –  Elongation –  Termination 3ʹ′ 5ʹ′ DNA 1 Initiation. After RNA polymerase binds to the promoter, the DNA strands unwind, and the polymerase initiates RNA synthesis at the start point on the template strand. 5ʹ′ 3ʹ′ Unwound DNA 3ʹ′ 5ʹ′ Template strand of DNA transcript 2 Elongation. The polymerase moves downstream, unwinding the DNA and elongating the RNA transcript 5ʹ′ → 3 ʹ′. In the wake of transcription, the DNA strands re-form a double helix. Rewound RNA RNA 5ʹ′ 3ʹ′ 3ʹ′ 5ʹ′ 3ʹ′ 5ʹ′ RNA transcript 3 Termination. Eventually, the RNA transcript is released, and the polymerase detaches from the DNA. 5ʹ′ 3ʹ′ 3ʹ′ 5ʹ′ 5ʹ′ Figure 17.7 Completed RNA transcript Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 3ʹ′
  • 26.
    Non-template strand of DNA Elongation RNAnucleotides RNA polymerase A 3ʹ′ T C C C A A T U 3ʹ′ end U 5ʹ′ C A T E A G G C A G T A Newly made RNA Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings C T Direction of transcription (“downstream”) 5ʹ′ G A Template strand of DNA
  • 27.
    RNA Polymerase Bindingand Initiation of Transcription •  Promoters signal the initiation of RNA synthesis •  Transcription factors –  Help eukaryotic RNA polymerase recognize promoter sequences TRANSCRIPTION RNA PROCESSING 1 Eukaryotic promoters DNA Pre-mRNA mRNA TRANSLATION Ribosome Polypeptide 5ʹ′ 3ʹ′ Promoter 3ʹ′ 5ʹ′ T A T A A AA AT A T T T T TATA box Start point Template DNA strand Several transcription factors 2 Transcription factors 5ʹ′ 3ʹ′ 3ʹ′ 5ʹ′ 3 Additional transcription factors RNA polymerase II 5ʹ′ 3ʹ′ Figure 17.8 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Transcription factors 3ʹ′ 5ʹ′ 5ʹ′ RNA transcript Transcription initiation complex
  • 28.
    Elongation of theRNA Strand •  As RNA polymerase moves along the DNA –  It continues to untwist the double helix, exposing about 10 to 20 DNA bases at a time for pairing with RNA nucleotides Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 29.
    Termination of Transcription • The mechanisms of termination –  Are different in prokaryotes and eukaryotes Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 30.
    •  Concept 17.3:Eukaryotic cells modify RNA after transcription •  Enzymes in the eukaryotic nucleus –  Modify pre-mRNA in specific ways before the genetic messages are dispatched to the cytoplasm Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 31.
    Alteration of mRNAEnds •  Each end of a pre-mRNA molecule is modified in a particular way –  The 5ʹ′ end receives a modified nucleotide cap –  The 3ʹ′ end gets a poly-A tail A modified guanine nucleotide added to the 5ʹ′ end TRANSCRIPTION RNA PROCESSING 50 to 250 adenine nucleotides added to the 3ʹ′ end DNA Pre-mRNA mRNA 5ʹ′ Protein-coding segment G P P P AAUAAA Ribosome TRANSLATION 5ʹ′ Cap Polypeptide Polyadenylation signal 5ʹ′ UTR Start codon Stop codon Figure 17.9 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 3ʹ′ UTR 3ʹ′ AAA…AAA Poly-A tail
  • 32.
    Split Genes andRNA Splicing •  RNA splicing –  Removes introns and joins exons TRANSCRIPTION RNA PROCESSING DNA Pre-mRNA 5ʹ′ Exon Intron Pre-mRNA 5ʹ′ Cap 30 31 1 Coding segment mRNA Ribosome Intron Exon Exon 3ʹ′ Poly-A tail 104 105 146 Introns cut out and exons spliced together TRANSLATION Polypeptide mRNA 5ʹ′ Cap 1 3ʹ′ UTR Figure 17.10 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Poly-A tail 146 3ʹ′ UTR
  • 33.
    •  Is carriedout by spliceosomes in some cases 5ʹ′ 1 RNA transcript (pre-mRNA) Intron Exon 1 Exon 2 Protein Other proteins snRNA snRNPs Spliceosome 2 5ʹ′ Spliceosome components 3 Figure 17.11 5ʹ′ mRNA Exon 1 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Exon 2 Cut-out intron
  • 34.
    Ribozymes •  Ribozymes –  Arecatalytic RNA molecules that function as enzymes and can splice RNA Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 35.
    The Functional andEvolutionary Importance of Introns •  The presence of introns –  Allows for alternative RNA splicing Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 36.
    •  Proteins oftenhave a modular architecture –  Consisting of discrete structural and functional regions called domains •  In many cases –  Different exons code for the different domains in a protein Gene DNA Exon 1 Intron Exon 2 Transcription RNA processing Intron Exon 3 Translation Domain 3 Domain 2 Domain 1 Figure 17.12 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Polypeptide
  • 37.
    •  Concept 17.4:Translation is the RNA-directed synthesis of a polypeptide: a closer look Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 38.
    Molecular Components ofTranslation •  A cell translates an mRNA message into protein –  With the help of transfer RNA (tRNA) Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 39.
    •  Translation: thebasic concept TRANSCRIPTION DNA mRNA Ribosome TRANSLATION Polypeptide Amino acids Polypeptide tRNA with amino acid Ribosome attached Ph e Gly tRNA A GC A A A U G G U U U G G C Codons 5ʹ′ Figure 17.13 mRNA Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Anticodon 3ʹ′
  • 40.
    •  Molecules oftRNA are not all identical –  Each carries a specific amino acid on one end –  Each has an anticodon on the other end Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 41.
    The Structure andFunction of Transfer RNA •  A tRNA molecule A –  Consists of a single RNA strand C that is only C about 80 nucleotides long –  Is roughly L-shaped 3ʹ′ A C C A 5ʹ′ C G G C C G U G U A A U A U U C UA C A C AG * G * G U G U * C C * * U C * * G AG C (a) Two-dimensional structure. The four base-paired regions and three G C U A loops are characteristic of all tRNAs, as is the base sequence of the * G amino acid attachment site at the 3ʹ′ end. The anticodon triplet is A A* unique to each tRNA type. (The asterisks mark bases that have been C U * chemically modified, a characteristic of tRNA.) A G A Figure 17.14a Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Amino acid attachment site Anticodon C U C G A G A G * * G A G G Hydrogen bonds
  • 42.
    5ʹ′ 3ʹ′ Amino acid attachment site Hydrogen bonds AAG 3ʹ′ Anticodon (b) Three-dimensional structure Figure 17.14b Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 5ʹ′ Anticodon (c) Symbol used in this book
  • 43.
    •  A specificenzyme called an aminoacyl-tRNA synthetase –  Joins each amino acid to the correct tRNA Amino acid Aminoacyl-tRNA synthetase (enzyme) 1 Active site binds the amino acid and ATP. P P P Adenosine ATP 2 ATP loses two P groups and joins amino acid as AMP. P Pyrophosphate Pi Phosphates 3 Appropriate tRNA covalently Bonds to amino Acid, displacing AMP. P Adenosine Pi Pi tRNA P Adenosine AMP 4 Activated amino acid is released by the enzyme. Figure 17.15 Aminoacyl tRNA (an “activated amino acid”) Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 44.
    Ribosomes •  Ribosomes –  Facilitatethe specific coupling of tRNA anticodons with mRNA codons during protein synthesis Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 45.
    •  The ribosomalsubunits –  Are constructed of proteins and RNA molecules named ribosomal RNA or rRNA DNA TRANSCRIPTION mRNA Ribosome TRANSLATION Polypeptide Growing polypeptide Exit tunnel tRNA molecules Large subunit E P A Small subunit 5ʹ′ mRNA Figure 17.16a 3ʹ′ (a) Computer model of functioning ribosome. This is a model of a bacterial ribosome, showing its overall shape. The eukaryotic ribosome is roughly similar. A ribosomal subunit is an aggregate of ribosomal RNA molecules and proteins. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 46.
    •  The ribosomehas three binding sites for tRNA –  The P site –  The A site –  The E site P site (Peptidyl-tRNA binding site) A site (AminoacyltRNA binding site) E site (Exit site) Large subunit E mRNA binding site Figure 17.16b P A Small subunit (b) Schematic model showing binding sites. A ribosome has an mRNA binding site and three tRNA binding sites, known as the A, P, and E sites. This schematic ribosome will appear in later diagrams. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 47.
    Amino end Growing polypeptide Nextamino acid to be added to polypeptide chain tRNA 3ʹ′ mRNA 5ʹ′ Codons (c) Schematic model with mRNA and tRNA. A tRNA fits into a binding site when its anticodon base-pairs with an mRNA codon. The P site holds the tRNA attached to the growing polypeptide. The A site holds the tRNA carrying the next amino acid to be added to the polypeptide chain. Discharged tRNA leaves via the E site. Figure 17.16c Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 48.
    Building a Polypeptide • We can divide translation into three stages –  Initiation –  Elongation –  Termination Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 49.
    Ribosome Association andInitiation of Translation •  The initiation stage of translation –  Brings together mRNA, tRNA bearing the first amino acid of the polypeptide, and two subunits of a ribosome P site 3ʹ′ U A C 5ʹ′ 5ʹ′ A U G 3ʹ′ Initiator tRNA GTP GDP E mRNA 5ʹ′ Start codon mRNA binding site Figure 17.17 3ʹ′ Small ribosomal subunit 1 A small ribosomal subunit binds to a molecule of mRNA. In a prokaryotic cell, the mRNA binding site on this subunit recognizes a specific nucleotide sequence on the mRNA just upstream of the start codon. An initiator tRNA, with the anticodon UAC, base-pairs with the start codon, AUG. This tRNA carries the amino acid methionine (Met). Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Large ribosomal subunit 5ʹ′ A 3ʹ′ Translation initiation complex 2 The arrival of a large ribosomal subunit completes the initiation complex. Proteins called initiation factors (not shown) are required to bring all the translation components together. GTP provides the energy for the assembly. The initiator tRNA is in the P site; the A site is available to the tRNA bearing the next amino acid.
  • 50.
    Elongation of thePolypeptide Chain •  In the elongation stage of translation –  Amino acids are added one by one to the preceding amino acid TRANSCRIPTION Amino end of polypeptide DNA mRNA Ribosome TRANSLATION Polypeptide mRNA Ribosome ready for next aminoacyl tRNA E 3ʹ′ P A site site 5ʹ′ 1 Codon recognition. The anticodon of an incoming aminoacyl tRNA base-pairs with the complementary mRNA codon in the A site. Hydrolysis of GTP increases the accuracy and efficiency of this step. 2 GTP 2 GDP E E P Figure 17.18 3 Translocation. The ribosome translocates the tRNA in the A site to the P site. The empty tRNA in the P site is moved to the E site, where it is released. The mRNA moves along with its bound tRNAs, bringing the next codon to be translated into the A site. P A GDP GTP Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings E P A A 2 Peptide bond formation. An rRNA molecule of the large subunit catalyzes the formation of a peptide bond between the new amino acid in the A site and the carboxyl end of the growing polypeptide in the P site. This step attaches the polypeptide to the tRNA in the A site.
  • 51.
    Termination of Translation • The final stage of translation is termination –  When the ribosome reaches a stop codon in the mRNA Release factor Free polypeptide 5ʹ′ 3ʹ′ 3ʹ′ 5ʹ′ 5ʹ′ 3ʹ′ Stop codon (UAG, UAA, or UGA) 1 When a ribosome reaches a stop 2 The release factor hydrolyzes 3 The two ribosomal subunits codon on mRNA, the A site of the the bond between the tRNA in and the other components of ribosome accepts a protein called the P site and the last amino the assembly dissociate. a release factor instead of tRNA. acid of the polypeptide chain. The polypeptide is thus freed from the ribosome. Figure 17.19 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 52.
    Polyribosomes •  A numberof ribosomes can translate a single mRNA molecule simultaneously –  Forming a polyribosome Completed polypeptide Growing polypeptides Incoming ribosomal subunits Start of mRNA (5ʹ′ end) Polyribosom End of mRNA (3ʹ′ end) (a) An mRNA molecule is generally translated simultaneously by several ribosomes in clusters called polyribosomes. e Ribosomes mRNA 0.1 µm Figure 17.20a, b (b) This micrograph shows a large polyribosome in a prokaryotic cell (TEM). Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 53.
    Completing and Targetingthe Functional Protein •  Polypeptide chains –  Undergo modifications after the translation process Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 54.
    Protein Folding andPost-Translational Modifications •  After translation –  Proteins may be modified in ways that affect their three-dimensional shape Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 55.
    Targeting Polypeptides toSpecific Locations •  Two populations of ribosomes are evident in cells –  Free and bound •  Free ribosomes in the cytosol –  Initiate the synthesis of all proteins Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 56.
    •  Proteins destinedfor the endomembrane system or for secretion –  Must be transported into the ER –  Have signal peptides to which a signalrecognition particle (SRP) binds, enabling the translation ribosome to bind to the ER Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 57.
    •  The signalmechanism for targeting proteins to the ER 1 Polypeptide synthesis begins on a free ribosome in the cytosol. 2 An SRP binds to the signal peptide, halting synthesis momentarily. 3 The SRP binds to a receptor protein in the ER membrane. This receptor is part of a protein complex (a translocation complex) that has a membrane pore and a signal-cleaving enzyme. 4 The SRP leaves, and 5 The signalthe polypeptide resumes cleaving growing, meanwhile enzyme translocating across the cuts off the membrane. (The signal signal peptide. peptide stays attached to the membrane.) 6 The rest of the completed polypeptide leaves the ribosome and folds into its final conformation. Ribosome mRNA Signal peptide Signalrecognition particle (SRP) SRP receptor CYTOSOL protein ERLUMEN Translocation complex Figure 17.21 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Signal peptide removed ER membrane Protein
  • 58.
    •  Concept 17.5:RNA plays multiple roles in the cell: a review •  RNA –  Can hydrogen-bond to other nucleic acid molecules –  Can assume a specific three-dimensional shape –  Has functional groups that allow it to act as a catalyst Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 59.
    •  Types ofRNA in a Eukaryotic Cell Table 17.1 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 60.
    •  Concept 17.6:Comparing gene expression in prokaryotes and eukaryotes reveals key differences •  Prokaryotic cells lack a nuclear envelope –  Allowing translation to begin while transcription is still in progress RNA polymerase DNA mRNA Polyribosome RNA polymerase Direction of transcription DNA Polyribosome Polypeptide (amino end) Ribosome Figure 17.22 0.25 µm mRNA (5ʹ′ end) Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 61.
    •  In aeukaryotic cell –  The nuclear envelope separates transcription from translation –  Extensive RNA processing occurs in the nucleus Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 62.
    •  Concept 17.7:Point mutations can affect protein structure and function •  Mutations –  Are changes in the genetic material of a cell •  Point mutations –  Are changes in just one base pair of a gene Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 63.
    •  The changeof a single nucleotide in the DNA’s template strand –  Leads to the production of an abnormal protein Wild-type hemoglobin DNA 3ʹ′ Mutant hemoglobin DNA C T T G U A 5ʹ′ In the DNA, the mutant template strand has an A where the wild-type template has a T. The mutant mRNA has a U instead of an A in one codon. 3ʹ′ 5ʹ′ T C A mRNA mRNA G A A 5ʹ′ 3ʹ′ 5ʹ′ 3ʹ′ Normal hemoglobin Sickle-cell hemoglobin Glu Val Figure 17.23 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings The mutant (sickle-cell) hemoglobin has a valine (Val) instead of a glutamic acid (Glu).
  • 64.
    Types of PointMutations •  Point mutations within a gene can be divided into two general categories –  Base-pair substitutions –  Base-pair insertions or deletions Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 65.
    Substitutions •  A base-pairsubstitution –  Is the replacement of one nucleotide and its partner with another pair of nucleotides –  Can cause missense or nonsense Wild type mRNA Protein 5ʹ′ A U G Met A A G U U U GG C U A A Lys Phe Gly 3ʹ′ Stop Amino end Carboxyl end Base-pair substitution No effect on amino acid sequence U instead of C A U G A A G U U U G G U U A A Met Lys Missense Phe Gly Stop A instead of G A U G A A G U U U A G U U A A Met Lys Phe Ser Stop Nonsense U instead of A A U G U A G U U U G G C U A A Figure 17.24 Met Stop Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 66.
    Insertions and Deletions • Insertions and deletions –  Are additions or losses of nucleotide pairs in a gene –  May produce frameshift mutations Wild type mRNA Protein 5ʹ′ A U GA A GU U U GG C U A A Met Lys Gly Phe Stop Amino end Carboxyl end Base-pair insertion or deletion Frameshift causing immediate nonsense Extra U AU GU A AG U U U G GC U A Met Stop Frameshift causing extensive missense U Missing A U G A A GU U G G C U A A Met Lys Leu Ala Insertion or deletion of 3 nucleotides: no frameshift but extra or missing amino acid A A G Missing A U G U U U G G C U A A Figure 17.25 Met Phe Gly Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Stop 3ʹ′
  • 67.
    Mutagens •  Spontaneous mutations – Can occur during DNA replication, recombination, or repair Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 68.
    •  Mutagens –  Arephysical or chemical agents that can cause mutations Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 69.
    What is agene? revisiting the question •  A gene –  Is a region of DNA whose final product is either a polypeptide or an RNA molecule Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings
  • 70.
    •  A summaryof transcription and translation in a eukaryotic cell DNA TRANSCRIPTION 1 RNA is transcribed from a DNA template. 3ʹ′ RNA transcript 5ʹ′ RNA polymerase RNA PROCESSING Exon 2 In eukaryotes, the RNA transcript (premRNA) is spliced and modified to produce mRNA, which moves from the nucleus to the cytoplasm. RNA transcript (pre-mRNA) Intron Aminoacyl-tRNA synthetase NUCLEUS Amino acid tRNA FORMATION OF INITIATION COMPLEX CYTOPLASM 3 After leaving the nucleus, mRNA attaches to the ribosome. mRNA AMINO ACID ACTIVATION 4 Each amino acid attaches to its proper tRNA with the help of a specific enzyme and ATP. Growing polypeptide Activated amino acid Ribosomal subunits 5ʹ′ TRANSLATION A succession of tRNAs add their amino acids to the polypeptide chain Anticodon as the mRNA is moved through the ribosome one codon at a time. (When completed, the polypeptide is released from the ribosome.) 5 E A AAA UG GU U U A U G Codon Figure 17.26 Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Ribosome