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DNARNAProteins Honors Biology REVIEW! What is DNA? Deoxyribonucleic Acid (DNA) Monomers made up of nucleotides: Nucleotides consist of: A five carbon sugar, deoxyribose o Four in it’s ring, one extending above the ring o Missing one oxygen when compared to ribose Phosphate group o Is the source of the “acid” in nucleic acid Nitrogenous base (Adenine, Guanine, Cytosine, Thymine) o A ring consisting of nitrogen and carbon atoms with various functional groups attached o Double ring= purines (A and G) o Single ring= pyrimidines (T and C) Double helix consists of: Sugar-phosphate backbone held by covalent bonds Nitrogen bases are hydrogen bonded together; A pairs with T and C pairs with G REVIEW! Nucleotides Protein synthesis: overview DNA inherited by an organism specifies traits by dictating the synthesis of proteins. However, a gene does not build a protein directly; it dispatches instruction in the form of RNA, which in turn programs protein synthesis. Message from DNA in the nucleus of the cell is sent on RNA to protein synthesis in the cytoplasm. Two main stages: Transcription Translation Protein Synthesis: Overview Two main stages: Transcription The transfer of genetic information from DNA into an RNA molecule Occurs in the eukaryotic cell nucleus RNA is transcribed from a template DNA strand Translation Transfer of the information in RNA into a protein. Transcription Details: 1. Initiation Promoter is the nucleotide sequence on DNA that marks where transcription of a gene begins and ends; “start” signal Promoter serves as a specific binding site for RNA polymerase and determines which of the two strands of the DNA double helix is used as the template. Specific nucleotide sequence at promoter is TATAAA Called the “TATA box”; located 25-35 base pairs before the transcription start site of a gene TATA box is able to define the direction of transcription and also indicates the DNA strand to be read Proteins called transcription factors can bind to the TATA box and recruit RNA polymerase; it has a regulatory function Note:TATA box is found upstream of start site and thus is NOT transcribed by RNA polymerase Transcription Elongation RNA elongates As RNA synthesis continues, the RNA strand peels away from its DNA template, allowing the two separated DNA strands to come back together in the region already transcribed. Transcription 3. Termination RNA polymerase reaches a sequence of bases in the DNA template called a terminator. Signals the end of the gene; at that point, the polymerase molecule detaches from the RNA molecule and the gene. mRNA (messenger RNA) or “transcript” exits the nucleus via the nuclear pores and enter the cytoplasm Transcription animation http://www- class.unl.edu/biochem/gp2/m_biology/animation/gene/ge ne_a2.html RNA processing Before mRNA leaves the nucleus, it is modified or processed. 1. addition of extra nucleotides to the ends of the transcript Include addition of a small cap (a single G nucleotide) at one end and a long tail (a chain of 50 to 250 A’s) at the other end Cap and tail facilitate the export of the mRNA from the nucleus, protecting the transcript from attack by cellular enzymes, and help ribosomes bind to the mRNA Cap and tail are NOT translated into protein. http://vcell.ndsu.edu/animations/mrnaprocessing/movie.htm RNA processing 2. RNA splicing Cutting-and-pasting process catalyzed by a complex of proteins and small RNA molecules, but sometime the RNA transcript itself catalyzes the process. Introns “intervening sequences”; internal noncoding regions Get removed from transcript before it leaves nucleus Exons Coding regions; parts of a gene that are expressed as amino acids Joined to produce an mRNA molecule with a continuous coding sequence Cap and tail are considered parts of the first and last exons, although are not translated into proteins. http://student.ccbcmd.edu/biotutorials/protsyn/exon.html RNA processing More animations http://www.pbs.org/wgbh/aso/tryit/dna/protein.html http://www.wisc- online.com/objects/index_tj.asp?objID=AP1302 Translation A typical gene consists or hundreds or thousands of nucleotides in a specific sequence, which get transcribed onto mRNA. Translation is the conversion of nucleic acid language into polypeptide language There are 20 different amino acids. A cell has a supply of amino acids in cytoplasm, either obtained by food or made from other chemicals. Flow of information from gene to protein is based on a triplet code: genetic instructions for the a.a. sequence of a polypeptide chain are written in DNA and mRNA as a series of three-base pairs, or codons. Translation- tRNA To convert the codons of nucleic acids on mRNA to the amino acids of proteins, a cell employs a molecular interpreter, called transfer RNA (tRNA) tRNA molecules are responsible for matching amino acids to the appropriate codons to form the new polypeptide. tRNA’s unique structure enables it to be able to: 1. pick up the appropriate amino acids 2. recognize the appropriate codons in the mRNA Translation- tRNA tRNA is made of a single strand of RNA consisting of about 80 nucleotides By twisting and folding upon itself, it forms several doublestranded regions in which short stretches of RNA base-pair with other stretches. at one end of the folded molecule contains a special triplet of bases called an anticodon. Complementary to a codon triplet on mRNA Anticodon recognizes a particular codon triplet on mRNA At the other end of the tRNA molecule is a site where an amino acid can attach. Translation- tRNA Translation- tRNA Each amino acid is joined to the correct tRNA by a specific enzyme. Each enzyme specifically binds one type of amino acid to all tRNA molecules that code for that amino acid, using a molecule of ATP as energy to drive the reaction. The resulting amino acid-tRNA complex can furnish its amino acid to a growing polypeptide chain. Translation- rRNA Ribosomal RNA (rRNA) Organelle in the cytoplasm that coordinates the functioning of mRNA and tRNA and actually makes polypeptides. Consists of two subunits: large and small Each ribosome has a binding site for mRNA, and three binding sites for tRNA. E site Removes tRNA from ribosome P site Holds the growing polypeptide A site Obtains new amino-acid-tRNA Ribosome holds tRNA and mRNA molecules close together, allowing the amino acids carried by the tRNA molecules to be connected into a polypeptide chain. Translation- Steps Can be divided into same three phases: initiation, elongation, and termination. 1. Initiation Brings together the mRNA, a tRNA bearing the first amino acid, and the two subunits of a ribosome. Role is to establish exactly where translation will begin, ensuring the mRNA codons are translated into the correct sequence of amino acids. Translation 1. Initiation (continued…) Two steps: 1. an mRNA binds to a small ribosomal subunit. A special initiator tRNA binds to the specific codon, called the start codon, where translation begins on mRNA. Initiator tRNA carries the amino acid Methionine (Met); its anticodon UAC binds to the start codon, AUG 2.A large ribosomal subunit binds to the smaller one, creating a function ribosome. The initiator tRNA fits into tRNA binding site (P site) on the ribosome. A site is vacant and ready for the next amino-acid carrying tRNA. 2. Elongation Once initiation is complete, amino acids are added one by one to the first amino acid. Each addition occurs in a three step process: 1. codon recognition The anticodon of an incoming tRNA carrying an amino acid, pairs with the mRNA codon in the A site of the ribosome 2. peptide bond formation Polypeptide separates from the tRNA to which it was bound (P site) and attaches by a peptide bond to the amino acid carried by the tRNA in the A site. The ribosome catalyzes formation of the bond. 3. translocation P site tRNA, moves to the E site and leaves the ribosome. The ribosome then translocates (moves) the tRNA in the A site, with its attached polypeptide, to the P site. Codon and anticodon remain bonded, and the mRNA and tRNA move as a unit Movement brings into the A site the next mRNA codon to be translated, and the process begins again at step 1. Termination Elongation continues until a stop codon reaches the ribosome’s A site. Stop codons- UAA, UAG, and UGA, do not code for amino acids but instead act as signal to stop translation. The completed polypeptide is released from the last tRNA and exits the ribosome, which then splits into its separate subunits. Translation Animation http://www- class.unl.edu/biochem/gp2/m_biology/animation/gene/ge ne_a3.html Polysome Several ribosomes can translate an mRNA at the same time, forming what is called a polysome. Peptide Bond Formation Free ribosomes vs. bound ribosomes Free ribosomes Found in cytoplasm Synthesize proteins for use primarily within the cell Bound ribosomes Found on rough ER Synthesize proteins primarily for secretion or for lysosomes Free ribosomes vs. bound ribosomes After protein synthesis… Each polypeptide coils and folds, assuming a 3-D shape, its tertiary structure. Several polypeptides may come together, forming a protein with quaternary structure. Overall significance: Process whereby genes control the structures and activities of cells The way genotypes determine phenotypes; proteins made from the original DNA nucleotides determine the appearance and capabilities of the cell and organism! Mutations Mutation is any change in the nucleotide sequence of DNA. Can involve large regions of a chromosome or just a single nucleotide pair, as in sickle cell disease In one of the two kinds of polypeptides in the hemoglobin protein, the sickle-cell individual has a single different amino acid. This small difference is caused by a change of a single nucleotide in the coding strand of DNA. Only ONE base pair! Mutations on DNA Two general categories: Base substitution Also known as a point mutation Replacement of one nucleotide with another. Depending on how the base substitution is translated, it can result in no change in the protein (due to redundancy of genetic code), an insignficant change, or a change that significantly affects the individual. Occasionally, it leads to an improved protein that enhances the success of the mutant organism and its descendants. More frequently, its harmful. o May cause changes in protein that prevent it from functionally normally. o If stop codon is a result of mutation and protein is shortened, it may not function at all. Mutations on DNA Base insertions or deletions Also known as frameshift mutation Often has a disastrous effect Adding or subtracting nucleotides may result in an alteration of the reading frame of the message all the nucleotides that are “downstream” of the insertion or deletion will be regrouped into different codons. Result will most likely by a nonfunctional polypeptide Mutations on DNA What causes mutations? Mutagenesis, or the production of mutations, can occur in a number of ways. Errors that occur during DNA replication or recombination are called spontaneous mutations. Mutagen, a physical or chemical agent that causes mutations Physical mutagen: high-energy radiation, such as X-rays and UV light Chemical mutagen: consists of chemicals that are similar to normal DNA bases pair incorrectly. Mutations on DNA Can also be helpful both in nature and in the laboratory. It is because of mutations that there is such a rich diversity of genes in the living world, that make evolution by natural selection possible. Also essential tools for geneticists. Whether naturally occurring or created in the laboratory, mutations create the different alleles needed for genetic research. Mutations- Chromosome Number Nondisjunction Members of a chromosome fail to separate. Can lead to an abnormal chromosome number in any sexually reproducing diploid organism. For example, if there is nondisjunction affecting human chromosome 21 during meiosis I, half the resulting gametes will carry an extra chromosome 21. Then, if one of these gametes unites with a normal gamete, trisomy 21 (Down Syndrome) will result. Mutations- Chromosome Number Mutations- Chromosome Structure Abnormalities in chromosome structure: Breakage of a chromosome can lead to a variety of rearrangements affecting the genes of that chromosome: 1. deletion: if a fragment of a chromosome is lost. Usually cause serious physical and mental problems. Deletion of chromosome 5 causes cri du chat syndrome: child is mentally retarded, has a small head with unusual facial features, and has a cry that sounds like the mewing of a distressed cats. Usually die in infancy or early childhood. Mutations- Chromosome Structure 2.duplication: if a fragment from one chromosome joins to a sister chromatid or homologous chromosome. 3.inversion: if a fragment reattaches to the original chromosome but in the reverse direction. Less likely than deletions or duplications to produce harmful effects, because all genes are still present in normal number 4. translocation: moves a segment from one chromosome to another nonhomologous chromosome Crossing over between nonhomologous chromosomes! Mutations- Chromosome Structure Karyotype The term karyotype refers to the chromosome complement of a cell or a whole organism. A karyotype is an ordered display of magnified images of an individual’s chromosomes arranged in pairs, starting with the longest. In particular, it shows the number, size, and shape of the chromosomes as seen during metaphase of mitosis. Chromosome numbers vary considerably among organisms and may differ between closely related species. Karytype Karyotypes are prepared from the nuclei of cultured white blood cells that are ‘frozen’ at the metaphase stage of mitosis. Shows the chromosomes condensed and doubled A photograph of the chromosomes is then cut up and the chromosomes are rearranged on a grid so that the homologous pairs are placed together. Homologous pairs are identified by their general shape, length, and the pattern of banding produced by a special staining technique. Karyotype Male karyotype Has 44 autosomes, a single X chromosome, and a Y chromosome (written as 44 + XY) Female karyotype Shows two X chromosomes (written as 44 + XX) Karyotype- Normal Karyotype- Abnormal