2021-Microbiology-Lecture-Notes-BIOL-3100-Microbial Genetics -9A PDF

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BIOL 3100 - Microbiology Lecture Notes

Microbial Genetics (DNA Replication & Protein Synthesis)

DNA, Genetic Engineering I. THE BASIS OF HEREDITY All information necessary for life is stored in an organism’s chromosomes, which are made up of DNA (exception: some viruses only have RNA). A chromosome is circular (prokaryotes) or linear (eukaryotes). Remember the organic molecules stuff we covered at the beginning of the semester? Nucleic acids (DNA & RNA) are made up of building blocks called nucleotides (more on structure below). In DNA, nucleotides are arranged in a twisted double chain called a helix. The particular nucleotide sequence spells out the “genetic code” that provides information for the synthesis of new DNA (DNA replication necessary for cell division) and for the synthesis of proteins. A typical prokaryotic cell contains a single circular chromosome. Bacteria may also have a small, circular piece of extrachromosomal DNA called a plasmid. Human body cells, examples of eukaryotic cells, have 46 linear chromosomes. A gene, the basic unit of heredity, is a liner sequence of DNA nucleotides that form a functional unit of the chromosome or plasmid. All information for the structure and function of an organism is coded in its genes. The information in specific gene is not always the same; different versions of the same gene are called alleles. Using humans as an example, the hair color gene is always found 1

BIOL 3100 - Microbiology Lecture Notes

at the same location on a chromosome, but the different versions or alleles that can exist for hair color are blond, brunette, red, etc. Because prokaryotes only have one chromosome, so they generally only have one allele for a particular gene. Many eukaryotes have 2 sets of chromosomes and thus 2 alleles of each gene, which may be the same or different. For example in humans, we have 46 chromosomes or 23 pair. In each pair, you get one chromosome from your mom and one for your dad. You may have a blonde hair allele from your mom and a dark hair allele from your dad (so you get dark hair since the dark hair allele is dominant). II. DNA STRUCTURE - THE WATSON-CRICK MODEL [DNA = deoxyribonucleic acid] In the Watson-Crick model, the DNA molecule is a double-stranded helix, shaped like a twisted "ladder." Remember that nucleic acids (DNA & RNA) are made up of building blocks called nucleotides. Each nucleotide is made up of a sugar, a phosphate, and a nitrogenous base. When we put these nucleotides together to build a DNA ladder, the sides of the ladder are composed of alternating phosphate groups & sugar molecules. The rungs of the ladder are made up of paired nitrogenous bases joined in the middle by hydrogen bonds. The nitrogenous bases are adenine, thymine, guanine, & cytosine; adenine always pairs with thymine (A-T or T-A) & guanine always pairs with cytosine (G-C or CG) [This is called complementary base pairing]. These 4 bases spell out the genetic message or code! 2

BIOL 3100 - Microbiology Lecture Notes

DNA enters into 2 kinds of reactions: 1.)

Replication - replicates the DNA before cell division, so that each new

daughter cell will receive a copy. 2.) Protein Synthesis (Gene Expression); 2 steps: transcription & translation

III.

DNA REPLICATION IN PROKARYOTES

[Remember bacteria have a circular chromosome.] Replication begins by an enzyme breaking the hydrogen bonds between the nitrogenous bases in the DNA molecule; the double stranded DNA molecule "unzips" down the middle, with the paired bases separating. As the 2 strands separate, they act as templates, each one directing the synthesis of a new complementary strand along its length.

If a nucleotide with thymine is present on the old strand, only a nucleotide with adenine can fit into place in the new strand; if a nucleotide with guanine is present on the old strand, only a nucleotide with cytosine can fit into place in the new strand, & so on. This is called complementary base pairing. DNA replication is called semiconservative replication since half of the original DNA molecule is conserved in each new DNA molecule. Like other biochemical reactions, DNA replication requires a number of different enzymes, each catalyzing a particular step in the process.

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BIOL 3100 - Microbiology Lecture Notes

IV. GENE EXPRESSION - PROTEIN SYNTHESIS A. FROM DNA TO PROTEIN: THE ROLE OF RNA By the 1940's biologists realized that all biochemical activities of the cell depend on specific enzymes; even the synthesis of enzymes depends on enzymes! Remember that the DNA molecule is a code that contains instructions for biological function & structure. Proteins (enzymes) carry out these instructions. The linear sequence of amino acids in a protein determines its 3-D structure & it is this 3-D structure that determines the protein's function. The big question was: How does the sequence of bases in DNA specify the sequence of amino acids in proteins? The search for the answer to this question led to the discovery of RNA (ribonucleic acid), which is similar in structure to DNA (deoxyribonucleic acid).

Three types of RNA: 1.

messenger RNA (mRNA) - single stranded; contains codons (3 base

codes); mRNA is constructed to copy or transcribe DNA sequences. 2.

ribosomal RNA (ribosomes!) (rRNA) - ribosomes "read" the code on the

mRNA molecule & send for the tRNA molecule carrying the appropriate amino acid. 3.

transfer RNA (tRNA) - clover leaf shaped; at least one kind for each of the

20 a. a. found in proteins; each tRNA molecule has 2 binding sites - one end, the anticodon (also a 3 base code), binds to the codon on the mRNA molecule; 4

BIOL 3100 - Microbiology Lecture Notes

the other end of the tRNA molecule binds to a specific amino acid; each tRNA & its anticodon are specific for an a. a.!! Differences between RNA & DNA: 1.

RNA nucleotides contain a different sugar than DNA nucleotides. (ribose

vs. deoxyribose). 2.

RNA is single stranded - DNA is double stranded.

3.

In RNA, uracil replaces thymine. There is no thyamine in RNA!!! But,

there is adenine.

B. TWO MAJOR EVENTS IN PROTEIN SYNTHESIS: 1. Transcription [mRNA copies or transcribes DNA sequences] This process is similar to what occurs in DNA replication. A segment of DNA uncoils unzips. Free RNA nucleotides, are then added one at a time to one end of the growing RNA chain. Cytosine in DNA dictates guanine in mRNA, guanine in DNA dictates cytosine in mRNA, adenine in DNA dictates uracil in mRNA, thymine in DNA dictates adenine in RNA. This complementary base pairing is just like what occurs in DNA replication. An enzyme catalyzes this process. After transcription the mRNA goes out in search of a ribosome. This mRNA molecule will now dictate the sequence of a. a. in a protein in the next step called translation.

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BIOL 3100 - Microbiology Lecture Notes

2.

Translation - actual synthesis of polypeptides or proteins; translate

information from one language (nucleic acid base code) into another language (amino acids); remember, the sequence of amino acids (the protein's primary structure) determines what the protein's 3-D globular structure is going to be & structure determines function. a.

Initiation - Begins when the ribosome attaches to the mRNA molecule,

reading its first or START codon. The first tRNA comes into place to pair with the initiator codon of mRNA (it occupies the peptide site in the ribosome). The START codon is AUG, which specifies the amino acid methionine. All newly synthesized polypeptides have to start with methionine.

b.

Elongation - The second codon of the mRNA molecule is then read and a

tRNA with an anticodon complementary to the second mRNA codon attaches to the mRNA molecule; with its a. a. this second tRNA molecule occupies the aminoacyl site of the ribosome. When both the P & A sites are occupied, an enzyme forges a peptide bond between the 2 a. a. & the first tRNA is released. The first tRNA cannot be released until this peptide bond is formed, as it will take its a. a. with it!! The second tRNA is then transferred from the A site to the P site & a third tRNA is brought into the A site. The ribosome continues to move down the mRNA molecule in this fashion, "reading" the codons on the mRNA molecule & adding amino acids to the growing polypeptide chain.

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BIOL 3100 - Microbiology Lecture Notes

c.

Termination - Toward the end of the coding sequence on the mRNA

molecule is a codon that serves as a termination signal. There are no tRNA anticodons to complementary base pair with this codon. Translation stops and the polypeptide chain is freed from the ribosome. Enzymes in the cell then degrade the mRNA strand.

[In eukaryotic cells, the polypeptide is taken up by the rough e.r. & is modified into a 3-D protein; the proteins are then packaged into transport vesicles (a piece of the e. r. pinches off around the protein); these vesicles transport the proteins to the golgi complex for further modification; the finished protein is pinched off in a piece of golgi membrane (another vesicle) and is transported to the part of the cell where it is needed. In the prokaryotic cell, none of these organelles exist, modification/processing of the polypeptide into a protein occurs in the cytoplasm.]

The genetic code. The mRNA codons for the 20 universal amino acids. See the table in your text of mRNA codons for the 20 amino acids. The 3-base codons are written to the left and the abbreviations of the amino acids they correspond to are written to the right. The amino acid abbreviations in the table are: Ala - alanine; Arg - arginine; Asn - apararagine; Asp - aspartamine; Cys - cysteine; Glu - glutamic acid; Gln glutamine;

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BIOL 3100 - Microbiology Lecture Notes

Gly - glycine; His - histidine; Ile - isoleucine; Leu - leucine; Lys - lysine; Met methionine; Phe - phenylalanine; Pro - proline; Ser - serine; Thr - threonine; Trp tryptophan; Tyr - tyrosine; Val - valine.

The code has been proven to be the same for all organisms from humans to bacteria - it's known as the universal genetic code.

Notice that most of the amino acids have more than one code (ex. Arg has 6 codes!). However, each code is specific for an amino acid (ex. UUU only codes for the amino acid Phe).

Three of the 64 codons do not specify amino acids. Instead they indicate STOP or termination of the translation process (they say "This is the end of the polypeptide.")

The START codon is AUG, which specifies the amino acid methionine. All newly synthesized polypeptides have to start with methionine. Since AUG is the only codon for methionine, when it occurs in the middle of a message, it is ignored as a START codon and is simply read as a methionine-specifying codon.

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