Biotechnology Unit 3: DNA to Proteins. From DNA to RNA

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1 From DNA to RNA Biotechnology Unit 3: DNA to Proteins I. After the discovery of the structure of DNA, the major question remaining was how does the stored in the 4 letter code of DNA direct the and of an entire organism II. The central dogma of molecular biology a. There are 2 general steps to the production of a from a i. Genes code for () ii. RNA codes for the () b. So, the flow of genetic information is from i. This is known as the of molecular biology c. Together, transcription and translation are the means by which cells their genetic information i. This is known as gene III. of DNA into RNA a. RNA is an produced from DNA in the process of protein i. RNA can have many forms and an incredibly wide range of within the cell b. Like DNA, RNA is a linear made up of four different linked together by phosphodiester bonds i. There are two major differences between DNA and RNA 1. The nucleotides in RNA are instead of deoxyribonucleotides a. They contain the 5-carbon sugar instead of deoxyribose 2. The bases involved in the ribonucleotides are A, C, G, and () a. Uracil replaces and complimentarily binds with with 2 hydrogen bonds just like thymine ii. There is also a fairly major difference between DNA and RNA 1. RNA is stranded instead of being a double helix like DNA a. This allows RNA to into many different shapes to accomplish many different c. All RNA in a cell is made by i. There are several similarities between transcription and DNA 1. A small portion of the DNA double helix is opened up and to expose the bases of the DNA 2. One of the two strands acts as a and ribonucleotides are added using base-pairing 3. The new RNA molecule (called a ) grows one nucleotide at a time

2 a. The enzyme responsible for reading the template and building the new RNA molecule is called and it reads from and builds from 5 to 3 like DNA polymerase i. RNA polymerase uses energy from ribonucleoside for energy just like DNA polymerase ii. Transcription does differ from DNA replication in some ways 1. The RNA transcript that is formed does not stay bonded to the template strand like DNA does a. The RNA ends up being stranded b. Just behind the region where nucleotides are being added, the RNA transcript is and the DNA double helix 2. Only one side of the DNA molecule acts a during transcription a. Both sides can be templates for different genes at different on the chromosome however b. The or strand is the strand that is by the RNA polymerase c. The other strand is called the or strand and is the for the RNA molecule 3. RNA molecules are only transcribed from regions of DNA a. This means that transcription only produces a segment of not more than a few base-pairs and not an entire DNA molecule 4. Transcription of the same gene can occur by several different RNA polymerases a. Because the transcripts are released almost, transcription can start over shortly after the RNA polymerase has moved on from the starting point b. It takes approximately to transcribe a molecule of RNA from a sized gene of 1500 nucleotide pairs i. As many as 15 RNA polymerases can be transcribing this gene at any one give time ii. This means over can be synthesized in a hour 5. RNA polymerase is very similar in its action as DNA polymerase but there are a couple key a. RNA polymerase polymerizes instead of deoxyribonucleotides b. RNA polymerase does not require RNA i. Because RNA does not store genetic information, the necessity of in transcription is lower than in DNA 1. RNA polymerases make approximately 1 mistake in every compared to the rate of one mistake in 10 7 nucleotides by DNA polymerase iii. Several types of RNA are in cells 1. The vast majority of genes in a cell s DNA code for the amino acid sequence of

3 a. The RNA molecules that are copied from these genes are called RNA () i. In, each mrna carries information from just one gene, coding for just one protein ii. In, several adjacent genes are often transcribed together into a single mrna which carries the information for several different proteins 2. In other genes, the final product of the gene is the itself a. These nonmessenger RNAs can serve a,, and components of cells and they are important to the translation of proteins and the regulation of gene expression b. RNA () forms the central core of the ribosomes where mrna is translated into proteins c. RNA () for the adaptors that hold the amino acids in place on the ribosome so they can be incorporated into the protein d. () serve as regulators of eukaryotic gene expression d. in DNA tell RNA polymerase where to start and finish i. Because of the fact that not all of the DNA of an organism for proteins and because not all of the genes are in all cells, there has to be a to tell RNA polymerase where to begin transcription ii. The beginning of a gene must be in order for transcription to begin iii. This process is different in and 1. In procaryotes, when RNA polymerase collides with a DNA molecule it attaches itself very and moves along the DNA until it finds a specific region called a a. At the promoter, the RNA polymerase binds and opens up the double helix in front of itself i. Promoter regions are which prevents transcription from occurring in any direction other than 5 to 3 b. The RNA polymerase then moves along the DNA double helix reading the strand and producing the RNA transcript until it reaches another specific region called the or stop site i. At the terminator site, the RNA polymerase from the DNA double helix c. The procaryotic RNA polymerase has a special subunit called a that helps to identify the promoter region i. Once the promoter is identified, the sigma factor itself from the RNA polymerase until transcription is and the polymerase has disengaged 2. In eucaryotes, the process is much more a. Unlike procaryotes, eucaryotes have different RNA polymerases i. RNA polymerase I and RNA polymerase III are responsible for transcribing,, and

4 ii. RNA polymerase II is responsible for transcribing b. Another difference from procaryotes is that eukaryotic RNA polymerase doesn t have a sigma factor and therefor is dependent upon a large group of accessory proteins called transcription which assemble at the before transcription can occur i. A group of assemble at a short section of DNA located 25 base-pairs upstream of the gene 1. This region is usually rich in A and T nucleotides and is called the ii. Once the RNA polymerase II is bound to the transcription factor complex, it is by adding a phosphate to its tail and that allows it to break away from the transcription factors and begin transcription 1. The transcription factors then release and are available to at another promoter region c. The of eukaryotic chromosomes also affects transcription because the individual genes are spread out considerably more than they are in procaryotes i. This allows for almost unlimited combinations of sequences for individual genes d. Finally, the of the eukaryotic chromosomes adds a level of complexity to transcription e. of transcripts i. In prokaryotes, the DNA is already in the and exposed to ribosomes 1. can attach to the 5 end of the transcript and begin the process of protein synthesis while transcription is still going ii. In eucaryotes, the DNA is inside the nucleus and is not exposed to the ribosomes until it leaves 1. Eucaryotic transcripts go through several steps of RNA before they can leave the nucleus a. These steps take place while transcription is occurring because the enzymes are attached to the of the RNA polymerase b. RNA involves adding methylated to the 5 end of the transcript c. involves cutting a small portion off of the 3 end of the transcript and then adds a few hundred creating a poly-a tail d. Both RNA capping and polyadenylation are thought to add and to help with the of the mrna out of the nucleus 2. Most eukaryotic genes are with noncoding regions that need to be from the transcript before translation can occur a. The noncoding regions are called and the coding regions are called i. The introns are usually much than the exons

5 IV. Biotechnology Unit 3: DNA to Proteins b. The length of a gene is transcribed including both introns and exons c. The introns need to be removed by RNA i. There are sequences within the introns that them as the regions that need to be cut out ii. The splicing of mrna is done by other RNA molecules called small nuclear RNAs () that combine with proteins to form small nuclear ribonucleoprotein particles () (pronounced snurps ) iii. The cutting follows what is known as a cut 3. splicing provides the ability for one mrna to be used to produce several different a. Splicing the RNA (keeping different exons) provides many different functional mrna molecules b. It is estimated that of human genes undergo alternative splicing iii. Once RNA processing and splicing is complete, the mrna must the nucleus to join with ribosomes to be translated into proteins is the process of converting the RNA sequence into a a. Translation involves a much more change than transcription i. The used to carry the code in DNA and RNA is based on 5 (but only 4 at a time) ii. The used to build proteins is composed of There cannot be a one to one relationship between RNA and proteins so we need what is referred to as the that is identical in nearly all life forms a. The mrna sequence is read in consecutive groups of 3 nucleotides called i. There are 4x4x4 = for codon combinations ii. Because there are only 20 amino acids, there is a built in meaning come amino acids are coded for by more than one codon b. This allows for three unique depending on where the reading begins i. There is a start codon that identifies the proper reading frame for each protein b. The rapid of mrna into a protein requires a molecular that moves along the RNA, captures and holds trna molecules, and links the incoming amino acids i. RNA (trnas) molecules match amino acids to the proper mrna codons 1. trnas are molecules roughly 80 nucleotides long that act as to bring in the correct amino acid that is coded for by each codon a. Each trna contains a region known as the which is complimentary to the codon on the mrna b. The 3 end of the trna holds a specific molecule c. trna molecules are said to be when the amino acid is attached

6 i. This is done by enzymes called aminoacyl-trna ii. Each amino acid has a synthetase that is responsible for connecting that amino acid to the trna molecules with the proper anticodons ii. are the protein manufacturing machine 1. They are made of over 50 different and several RNA (rrna) molecules 2. Typical cells have of individual ribosomes a. Both eucaryotic and procaryotic ribosomes are similar and are made up of a subunit and a subunit i. The small subunit pairs the to the mrna codons ii. The large subunit links the into the polypeptide chain b. The ribosome attaches near the of the mrna and then pulls the mrna through while it the appropriate amino acids i. Eucaryotic ribosomes can add about per second ii. Procaryotic ribosomes can combine up to per second 3. Along with the binding site for the mrna, the ribosome has sites for trnas a. The is where the new trna comes in and binds to the anticodon and brings in the new amino acid b. The is where the previous trna is held until the new amino acid can be linked onto the chain c. The holds the trna until it is ejected d. The mrna moves through these sites until the polypeptide chain is complete 4. Codons in mrna signal where to and protein synthesis a. Translation always begins with the codon i. The trna carries the amino acid methionine and attaches to the AUG codon ii. This initiator trna is different from the normal carrying trna iii. The ribosomal subunit attaches to the 5 end of them RNA (signaled by the 5 cap) and then along until it finds an AUG b. It then reads along adding all the necessary amino acids until it reaches a codon i.,, and are the stop codons ii. Stop codons do not correspond to a trna and instead signal the ribosome to stop iii. factors (proteins) bind to the stop codon in the A-site and cause the and the to be released from the mrna

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9 Overview of Gene Expression Biotechnology Unit 3: DNA to Proteins I. Every cell in an organism has a complete set of the for that organism but not all of the are needed within the cell a. Different cell types produce different sets of b. The of each protein that the cell needs is different at different times c. The control of gene is how the cell deals with these issues i. Cells can change the expression of genes in response to signals II. Gene expression can be at many different levels or step within the from DNA to RNA to protein a. control i. Transcription is controlled by binding to DNA sequences 1. Remember that in order for transcription to begin, RNA must attach and itself at the promoter region of the gene 2. The simplest regulatory sequences are only long a. These are most common in 3. In eucaryotes, the regulatory sequences can be up to nucleotide pairs and can interact with a wide variety of signals 4. Transcription are proteins that bind to specific regulatory DNA sequences to act as a for the control of transcription a. Different proteins recognize different based on the shape of the protein ii. In procaryotes, control the expression of groups of genes 1. There is a region at the start of the operon called the which is where transcription of the entire group of genes will begin 2. Operators are controlled by and a. A repressor is a that when bound to the operator does not allow transcription to begin by blocking the of RNA polymerase b. An activator is a protein that allows transcription to occur by allowing for attachment of the DNA polymerase c. Some operons use repressors and activators 3. Operon a. operon i. This operon uses a and it controls a group of five genes that are necessary for the production of the amino acid tryptophan ii. When tryptophan is in in the cell, it can bind to the repressor protein which will then bind to the operon and transcription iii. When tryptophan is in low concentration, it cannot bind to the repressor protein and the repressor protein then cannot bind to the operator and transcription will occur for the production of tryptophan Essential Cell Biology Chapter 8

10 b. Lac operon i. This operon uses both a and an to control the transcription of several genes used to break down which is a food source for bacteria ii. is the bacterial cells first choice for food so when glucose is presence the Lac operon is switched off because it is lacking a necessary protein that is only available when glucose is absent iii. The operon is also switched off by a repressor protein that can only be removed when is present iv. The only time the Lac operon is active, is when both the activator is bound ( not present) and the repressor is not bound ( is present) Essential Cell Biology Chapter 8

11 iii. also use transcription regulators (both repressors and activators) but they do not use to control groups of genes 1. These sites are called 2. Eucaryotic transcription regulators can work from a a. The binding of activators or repressors in eucaryotes does not need to be to the gene of interest or even be from it b. In many instances the transcription regulators are long distances away but there is a that occurs to allow for the initiation of transcription b. -transcriptional control i. These are controls that occur the RNA polymerase has already started synthesizing the mrna ii. Some mrna molecules contain, short RNA segments that can change their conformation when certain chemicals are present and therefore can control their own expression 1. There are no involved in this type of regulation iii. Controlling 1. Repressor proteins can bind onto the in eucaryotic mrna and regulate when the ribosomes can attach to carry out translation 2. RNA (RNAi) is one way organisms can control translation a. Many have double stranded RNA (dsrna) i. completely or just parts ii. As a defense against these viruses, cells have the ability to dsrna and trigger the RNAi response b. A nuclease named recognizes and cuts dsrna into small pieces called RNAs (sirna) that are usually around 20 base pairs long i. This is a highly conserved nuclease found in organisms 1. It probably evolved to the divergence of plants, animals, and fungi 2. Recently, a similar enzyme has been found in ii. sirnas from dicer combine with a group of proteins to form a complex known as (RNA-induced silencing complex) 1. The sirna is into a single strand a. In some systems there is selection of strands but in most both an and are used 2. One of the key proteins is a protein called iii. The RISC complex then uses the single stranded sirna to bind to the mrna iv. Slicer then the mrna and the cell the sliced transcripts c. Another type of double stranded RNA exists that is found naturally in many cells called (mirna) i. These are expressed from regions of the genome and make natural double stranded RNA Essential Cell Biology Chapter 8

12 ii. mirnas are also recognized by dicer and target other genes as a form of controlling gene expression without cleavage iii. mirnas do not induce cleavage of the mrna because they are not complimentary 1. Instead they are involved in translational 2. Specific type of mirna called a small temporal RNA (strna) iv. Dicer interacts with the strna or other mrna and then they enter the RISC complex and bind to mrna but translation instead of cutting v. There is recognition so slicing does not occur vi. This allows for the same mirna to target several different or of genes Essential Cell Biology Chapter 8