jimtrue.com : school : BSC2010 : CH 17: From Gene to Protein

Posted by Jim True on April 20, 2004 6:45 AM. Last Updated October 22, 2006 9:23 PM

Disclaimer for all material noted here is at the bottom of this web page.

CH 17: From Gene to Protein

Genes and Protein

• Ideally it was thought that genes control the pheotypic expression by specifying enzymes that carry out reactions to produce the phenotype. Gene expression can also be called 'protein synthesis'.
• While many of the direct products of genetic expression are enzymes, genes also produce a wide variety of other polypeptides, plus RNA.
• Thus, our current hypothesis of the direct expression of genes is one gene -- one polypeptide.

RNA

• Differences between RNA and DNA:
  • 5 carbon sugar is ribose.
  • Pyrimidine uracil replaces thymine.
  • Is a single stranded molecule.
• There are three basic categories of RNA:
  1. Ribosomal RNA (rRNA) -- Found in ribosomes. They function as the site in a ribosome where polypeptides are assembled, ie. protein synthesis.
  2. Transfer RNA (tRNA) -- These are found as free molecules in the cytoplasm. There are over 40 different tRNA molecules in human cells.
     • Their function is to bring amino acids to the ribosome and transfer them to the ribosome to be added to the polypeptide (protein) being synthesized.
  3. Messenger (mRNA) -- Is created in the nucleus (eukaryote) or cytoplasm (prokaryote) as a copy of a gene.
     • The process of its formation is called transcription. (to copy across)
     • It moves from the nucleus to the ribosomes, either those in the nucleus, or those in the cytoplasm (at rough ER).
     • The instructions from mRNA are then used to synthesize a particular protein in the process of translation.
• Protein synthesis can also be referred to as "gene expression" since all genes "code" for the production of a polypeptide, protein, enzyme or RNA molecule.
• In prokaryotic cells, transcription and translation are directly linked together, as there is no nuclear membrane present.
• In eukaryotic cells, there are intermediate processes that occur between the two. (Figure 17.2, p.306)

The Genetic "Code"

• Since it is known that genes code for proteins, then what must be specified by the nucleotides are the individual amino acids.
• How do nucleotides provide information about specific amino acids?
• There are 20 known amino acids.
• Could each of the 4 single nitrogen bases taken one at a time be used to specify an amino acid?
• 4¹ = 4, so no way this would work.
• how about 2 bases in any combination (eg AT, GG, etc)?
• 4² = 16, also does not work. How about 3 at a time in any combination (eg AGG)?
• 4³ = 64. Yes, this works.
• There are 64 triplet base combinations that specify particular amino acids. These triplets are called codons.
• But there are only 20 amino acids! What about the other 44 codons?
• Many amino acids have multiple codons.
• The number of codons that specify a particular amino acid range from 1 to 6.
• One codon, AUG, specifies the amino acid methionine. The VERY FIRST time it is encountered in an mRNA by the rRNA "reader", AUG also specifies the START of protein synthesis.
• Three codons are called nonsense, terminal, or STOP codons..
• When one of these 3 (UAA, UAG, UGA) is encountered, it signifies the end of the synthesis of a particular protein. (figures 17.3, p.307, 17.4, p.308).

Transcription

• Transcription ("copy across") -- Occurs at the DNA molecule in the nucleus of eukaryotes or in the cytoplasm of prokaryotes (figure 17.6, p.309)
• It is the formation of a mRNA molecule as a copy of one gene on one strand of DNA.
• Transcription directly results in a functional mRNA in prokaryotes, but in eukaryotes, it is a two-step process.
• The first product is a primary transcript or pre-mRNa, which is then processed to produce the functional mRNA used in translation. (Rough draft).
• Transcription consists of three stages:
  1. Initiation -- The enzyme RNA polymerase attaches to a region of the DNA molecule and unwinds it.
     • On one strand an embedded series of nucleotides identify it as the template (AKA sense) strand, the strand that is copied.
     • Embedded in the sense strand are nucleotides that act as PROMOTER (Start) and TERMINATOR (Stop) signals to the RNA polymerase.
     • The enzyme will "read" the nitrogenous bases in between the start and stop nucleotides (transcription unit).
     • Like DNA polymerase, RNa polymerase can only "read" from 3'--5' along the DNa strand and construct mRNA 5'--3'.
     • It attaches to the template strand by means of transcription factors, proteins that help it to bind to the template strand.
  2. Elongation -- RNA polymerase untwists DNA as it moves along the transcription unit.
     • Unlike DNA synthesis, the opened part of the strand immediately rewinds behind the RNa polymerase.
     • The pre-mRNA strand (or mRNA strand in prokaryotes) trails out into the nucleoplasm (or cytoplasm). (figure 17.6, p.309)
  3. Termination -- in Prokaryotes, the mRNA is released as the polymerase reaches the termination signal.
     • In eukaryotes, the polymerase continues past the termination sequence for several hundred nucleotides, but cuts the pre-mRNA free about 35 nucleotides past the terminator, producing the primary transcript.
     • The pre-mRNA will then undergo processing.

RNA Processing

• The pre-mRNA is processed at each end as well as within itself.
• At the 5' end, a modified guanine molecule is attached, forming a 5' cap.
• This serves to protect the mRNA from being broken down, and as an attachment point for the ribosome during translation.
• At the 3' end, a series of 50-250 adenine molecules are attached, forming the poly (A) tail, also helping in protection of mRNA and for attachment.
• When first formed, the pre-mRNA molecule is FAR larger than the number of nucleotides needed to code for the specific protein.
• This is because there are numerous regions that are not translatied.
• These are referred to as intervening sequences, or introns.
• The translatable portions of the pre-mRNA are known as expressed sequences, or exons.
• Pre-mRNA is processed by a complex of proteins and RNA that strip out the introns and join the exons together, a process called gene splicing.
• When completed, the functional mRNa moves to the ribosomes of the nucleus or rough ER, depending on where protein synthesis is to take place.

Translation

• Translation-- ("to convert from one form to another") -- Occurs in either the nucleoplasm at nuclear ribosomes or in cytoplasm at the rough ER.
• Present in the cytoplasm of the cell are the 20 amino acids, either synthesized within the cells or brought in from food (essential amino acids).
• Also present are transfer RNA molecules which act to "interpret" the mRNA.
• Each tRNA has three loops and a "tail" region to which specific amino acids attach. (Figure 17.12, p.315)
• On the loop opposite the tail is an exposed region of three bases called the anticodon.
• The anticodon match to the codon ensures that the correct amino acid is brought to the ribosome. The CODON codes for the amino acid, not the anticodon.
• The ribosome is formed of a complex of proteins and rRNA, and actually has two subunits, one large and one small. (figure 17.15, p.316)
• The large unit contains two depressions the P site and the A Site, and a tubular channel called the E site.
• The P site is where the tRNA and its amino acid are detached from each other.
• The A site is where the next codon is "read" and the next tRNA brings the next amino acid in the series.
• the E Site is where the tRNa molecules exit after releasing their amino acids.
• There are three stages to translation:
  1. Initiation -- A small ribosomal subunit attaches to the 5' end of the mRNA.
     • The initiation codon is AUG, and tRNA bearing methionine moves to the small subunit.
     • A large ribosomal subunit then attaches, locking the tRNA and methionine into the P Site. (for initiation ONLY locks down at the P site, after iniation, process occurs at A (amino acid read) to P (peptide bond formed) to E (eject tRNA)).
     • This is the translation initiation complex, and is assisted by protein initiation factors.
  2. Elongation -- Once the initiation complex is formed, the next codon is read at the A site, and the appropriate tRNA brings the next amino acid.
     • If this amino acid is accepted, a peptide bond is formed by rRNA between the amino acid at the A site and the one at the P site.
     • The tRNA at the P site is released, and the second amino acid now shifts to the P site, and the third codon is read at the A site.
     • This continues with peptide bonds forming to produce a lengthening string of amino acids, until a stop codon is read.
  3. Termination -- When a stop codon is read, a protein release factor binds to the stop codon in the A site. (figure 17.19, p.319)
     • The polypeptide is released from the ribosome and the ribosome disassembles into the large and small subunits.
     • It is possible for numerous ribosomes to attach to the same mRNA, once each preceding ribosome has moved past the initiation codon.
     • This results in polyribosomes, which basically produce numerous copies of the same polypeptide very quickly from the same set of instructions. Produce something that is needed in quantity without having to produce multiple mRNA's.

Mutation

• Mutation -- any alteration of a nucleotide sequence and therefore, a gene.
• In chapter 15, we discussed mutations due to large segments of chromosomes being rearranged.
• Point mutations are those that typically involve only a single base pair. If not corrected, these mutations persist in all copies of the gene.
• There are two categories of point mutations. (figure 17.24, p323)
  1. Substitution -- replacement of one base pair with another.
     • Some of these may be silent mutations, ie, result in no changes because the substitution STILL calls for the same aminio acid, eg if CUU mutated to CUA.
     • Base pair substitutions may alter the nature of the protein. In some cases, this can be actually BENEFICIAL, although usually it is HARMFUL.
     • Substitutions that still code for an amino acid, although not necessarily the same one, are missense mutations
     • Alterations that result in the formation of a stop codon are called nonsense mutations, because a non-functional protein will result.
  2. Frame Shift Mutations.
     • Insertion -- Addition of a nucleotide pair.
     • Deletion -- Removal of a nucleotide pair.
     • In either case, the sequence of codons in mRNA will change.
     • This causes a frameshift mutation, in which case the protein can be much longer than it is supposed to be (another missense as frameshift alters the stop codon to a later position), or shorter (another nonsense as frameshift alters the stop codon to an earlier position.
• Mutations can occur naturally, but can also be generated by exposure to mutagens.
• Mutagen -- any substance or condition that induces mutation. "agent of mutation".
• Includes both radiation sources, as well as chemical compounds.

Disclaimer: These are MY notes taken from classroom lectures while I'm in the classroom. While I'm perfectly happy to share my notes with my classmates and I know I take very good notes, you should still make every effort to attend the class and TAKE YOUR OWN NOTES. I will not transcribe everything the instructor says in the classroom, and I will NEVER post pre-exam reviews. My notes will not replace the value of actually attending class and taking your own class notes.I also cannot attest to their accuracy, other than they are what was provided in the lecture; you should not reference my notes as "expert opionion" by any means, and if you notice an error or omission, please do me the favor of e-mailing me with the correction and I will re-post my notes. End of Disclaimer.