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From Genes to Proteins


From Genes to Proteins
Although DNA determines the type of biochemical product needed by the cell and directs its synthesis, it is RNA through the process of translation, which is responsible for the actual assembly of the products.


RNA Structure and Function
RNA, like DNA, is a large molecule made up of a long string of nucleotides. However, it differs from DNA in three aspects of its structure. First, RNA is a single-stranded rather than a double-stranded molecule. Second, the sugar in each nucleotide of RNA is ribose instead of deoxyribose. Third, the pyrimidine base thymine in DNA is replaced by uracil in RNA.
Cells contain three types of RNA: messenger RNA (mRNA), ribosomal RNA (rRNA), and transfer RNA (tRNA). All three types of RNA are synthesized in the nucleus by RNA polymerase enzymes and then moved into the cytoplasm, where protein synthesis takes place. Messenger RNA carries the instructions for protein synthesis, obtained from the DNA molecule, into the cytoplasm. Transfer RNA reads the instructions and delivers the appropriate amino acids to the ribosome, where ribosomal RNA translates the instructions and provides the machinery needed for protein synthesis.

Messenger RNA. Messenger RNA is the template for protein synthesis. It is a long molecule containing several hundred to several thousand nucleotides. Each group of three nucleotides forms a codon that is exactly complementary to a nucleotide triplet of the DNA molecule. Messenger RNA is formed by a process called transcription.  In this process, the weak hydro- gen bonds of DNA are broken so that free RNA nucleotides can pair with their exposed DNA counterparts on the meaningful strand of the DNA molecule (see Fig. 6.4). As with the base pairing of the DNA strands, complementary RNA bases pair with the DNA bases. In RNA, uracil (U) replaces thymine and pairs with adenine. As with DNA, guanine pairs with cytosine.

The DNA helix and transcription of messenger RNA (mRNA). The DNA helix unwinds and a new mRNA strand is built on the template strand of the DNA

Ribosomal RNA. The ribosome is the physical structure in the cytoplasm where protein synthesis takes place. Ribosomal RNA forms 60% of the ribosome, with the remainder of the ribosome composed of the structural proteins and enzymes needed for protein synthesis. As with the other types of RNA, rRNA is synthesized in the nucleus. Unlike the two other types of RNA, rRNA is produced in a specialized nuclear structure called the nucleolus. The formed rRNA combines with ribosomal proteins in the nucleus to produce the ribosome, which is then transported into the cytoplasm. On reaching the cytoplasm, most ribosomes become attached to the endoplasmic reticulum and begin the task of protein synthesis.

Transfer RNA. Transfer RNA is a clover-shaped molecule containing only 80 nucleotides, making it the smallest RNA molecule. Its function is to deliver the activated form of an amino acid to the protein that is being synthesized in the ribosomes. At least 20 different types of tRNA are known, each of which recognizes and binds to only one type of amino acid. Each tRNA molecule has two recognition sites: the first is complementary for the mRNA codon and the second for the amino acid itself. Each type of tRNA carries its own specific amino acid to the ribosomes, where protein synthesis is taking place; there it recognizes the appropriate codon on the mRNA and delivers the amino acid to the newly forming protein molecule.

Transcription
Transcription occurs in the cell nucleus and involves the syn- thesis of RNA from a DNA template (Fig. 6.4). Genes are transcribed by enzymes called RNA polymerases that generate a single-stranded RNA identical in sequence (with the exception of U in place of T) to one of the strands of DNA. It is initiated by the assembly of a transcription complex composed of RNA polymerase and other associated factors. This complex binds to the double-stranded DNA at a specific site called the promoter region. Within the promoter region, the so-called TATA box is located. The TATA box contains the crucial thy- mine–adenine–thymine–adenine (TATA) nucleotide sequence that RNA polymerase recognizes and binds to. This binding also requires transcription factors, a transcription initiation site, and other proteins. Transcription continues to copy the meaningful strand into a single strand of RNA as it travels along the length of the gene, stopping only when it reaches a termination site with a stop codon. On reaching the stop signal, the RNA polymerase enzyme leaves the gene and releases the RNA strand. The RNA strand then is processed.
Processing involves the addition of certain nucleic acids at the ends of the RNA strand and cutting and splicing of certain internal sequences. Splicing involves the removal of stretches of RNA. Because of the splicing process, the final mRNA sequence is different from the original DNA template. The retained protein-coding regions of the mRNA sequences are called exons and the regions between exons are called introns. The functions of the introns are unknown. They are thought to be involved in the activation or deactivation of genes during various stages of development.
Splicing permits a cell to produce a variety of mRNA molecules from a single gene. By varying the splicing segments of the initial mRNA, different mRNA molecules are formed. For example, in a muscle cell, the original tropomyosin mRNA is spliced in as many as 10 different ways, yielding distinctly different protein products. This permits different proteins to be expressed from a single gene and reduces how much DNA must be contained in the genome.

Translation
Translation occurs in the cytoplasm of the cell and involves the synthesis of a protein using its mRNA template. Proteins are made from a standard set of amino acids, which are joined end to end to form the long polypeptide chains of protein molecules. Each polypeptide chain may have as many as 100 to more than 300 amino acids in it. Besides rRNA, translation requires the coordinated actions of mRNA and tRNA (Fig. 6.5). Each of the 20 different tRNA molecules transports its specific amino acid to the ribosome for incorporation into the developing protein molecule. Messenger RNA provides the information needed for placing the amino acids in their proper order for each specific type of protein. During protein synthesis, mRNA contacts and passes through the ribosome, during which it “reads” the directions for protein synthesis. As mRNA passes through the ribosome, tRNA delivers the appropriate amino acids for attachment to the growing polypeptide chain. The long mRNA molecule usually travels through and directs protein synthesis in more than one ribosome at a time. After the first part of the mRNA is read by the first ribosome, it moves onto a second and a third. As a result, ribosomes that are actively involved in protein synthesis are often found in clusters called polyribosomes.
Protein synthesis. A messenger RNA (mRNA) strand is shown moving along a small ribosomal subunit in the cytoplasm.

The process of translation is not over when the genetic code has been used to create the sequence of amino acids that constitute a protein. To be useful to a cell, this new polypeptide chain must fold up into its unique three-dimensional conformation. The folding of many proteins is made more efficient by special classes of proteins called molecular chaperones. Typically the function of a chaperone is to assist a newly synthesized polypeptide chain to attain a functional conformation as a new protein and then to assist the protein’s arrival at the site in the cell where the protein carries out its function. Molecular chaperones also assist in preventing the misfolding of existing proteins. Disruption of chaperoning mechanisms causes intracellular molecules to become denatured and insoluble. These denatured proteins tend to stick to one another, precipitate, and form inclusion bodies. The development of inclusion bodies is a common pathologic process in Parkinson, Alzheimer, and Huntington diseases.
A newly synthesized polypeptide chain may also need to combine with one or more polypeptide chains from the same or an adjacent chromosome, bind small cofactors for its activity, or undergo appropriate enzyme modification. During the posttranslation process, two or more peptide chains may combine to form a single product. For example, two α-globin chains and two β-globin chains combine to form the α2β2-hemoglobin molecule. The protein products may also be modified chemically by the addition of various types of functional groups. For example, fatty acids may be added, providing hydrophobic regions for attachment to cell membranes. Other modifications may involve cleavage of the protein, either to remove a specific amino acid sequence or to split the molecule into smaller chains. As an example, the two chains that make up the circulating active insulin molecule, one containing 21 and the other 30 amino acids, were originally part of an 82-amino-acid proinsulin molecule.