Microbial Genetics Part 1

1. Microbial Genetics Part 1 Genetics can be a challenge to understand. Use the McGraw Hill website to supplement this lecture. www.mhhe.com/cowan1 Please do not spend any time studying the Lactose Operon.

2. Over view of Microbial Genetics • 1. Replication of DNA: occurs before each cell division is complete. • 2. Transcripton: DNA is converted to RNA and occurs to carry on life processes. • 3. Translation: RNA is converted to protein (enzymes). • 4. Genetic Transfer and Recombination: How do we get genetic diversity (antibiotic resistance for example) within the microbial community?

3. Replication • Each organism has its own genome. A genome is all of the cell’s genetic information. Included in the genome are chromosomes and plasmids, as well as other DNA that is sometimes found within microbes. • Chromosomes are structures made up of DNA that carry hereditary information. (Remember that they are circular in bacteria.) • Genes are segments of DNA within chromosomes, that code for functional products. For example, the insulin gene codes for the final insulin product. • Each organism has a genotype and a phenotype. • The Genotype is the genetic make up of the organism. In other words, all the genes that it has. • The Phenotype is the manifest characteristics due to the genes it has. In other words, do you have blue eyes or green eyes?

4. DNA is composed of 4 nucleotides: Adenine (A), Thymine (T), Cytosine (C), and Guanine (G). • AT are complementary base pairs and CG are complementary base pairs. • To give you an idea of the size of DNA in bacteria, E-coli has 4 million bps. = 4,000 Kb in its one chromosome. • Each chromosome consists of 2 strands of DNA bases bound together. They are not identical to each other but are complementary to each other. • In order to know which end is which, they are labeled 3’ and 5’. (See Fig. 9.4 in your textbook) • Note that the 3’side of the nucleotide has a different chemical group from that of the 5’ side.

5. Steps of Replication • 1. DNA is partially unwound with the help of an enzyme called a helicase. The point where the helicase pauses the unwinding is called the replication fork. • 2. A molecule, called an RNA primer, is place on the DNA to help the nucleotides begin to bind. The complementary bases are then added to the template (parent) strand using an enzyme called polymerase. • DNA can only replicate in the 5’to 3’ direction. The reason is because the chemical group on 3’ side of the nucleotide acts like a hand that can grab onto the next nucleotide on its 5’side. • Since the DNA strands are complementary, (also called antiparallel) only one strand can replicate quickly and easily in the 5’ to 3’ direction. This is called the “leading strand”. • .

6. A little help is needed for the opposite strand so that it too can be replicated. • The first step consists of multiple RNA primers placed along the template strand. These primers provide the necessary hand for the nucleotides to grab onto. Then they can replicate the strand 5’ to 3’ for a short distance. These fragments of DNA are called Okazaki Fragments. • Once the strand has been replicated, the RNA primers are cut out and replaced by the missing nucleotides. This strand is called the “lagging strand” because it takes longer for it to be replicated. • 3. Once the strands are replicated up to the replication fork, the helicase unwinds the DNA some more and the replication fork moves down strand to a new location. • 4. The newly replicated DNA rewinds. One new strand winds together with one old strand. • This process of replication is called Semi-conservative Replication because one half of the template DNA is kept with one half of the new DNA.

7. In Prokaryotes, replication begins at a specific site in the chromosome called the origin of replication. • Because prokaryotes have a circular chromosome replication can proceed bi-directionally or rolling circle. • Bi-directional means that replication starts at the origin of replication and proceeds right and left on both strands. See Fig. 9.6 • Rolling Circle means that replication only occurs right or left from the origin of replication but as it proceeds, the DNA comes off of the chromosome in a motion similar to a tape dispenser. When replication is complete, the new chromosome is stitched into a circle using an enzyme called ligase. • The replication speed for E.coli is estimated to be 1000 nucleotides/sec.

8. Transcription • Formation of RNA from DNA • The nucleotides are essentially the same as the DNA nucleotides. The main difference is that Uracil (U) replaces Thymine (T) in RNA. In other words, anytime T would have been placed in the new strand, U is put in that spot instead. • RNA is single stranded, not double stranded. • 3 types of RNA are formed: • mRNA, messenger RNA is the template for protein synthesis. • tRNA, transfer RNAs are taxis for amino acids during protein synthesis. • rRNA, ribosomal RNAs are the site of protein synthesis. They put the protein together.

9. 1. RNA synthesis begins at a place on DNA called the promoter. • 2. DNA is unwound. • 3. A primer is put in place and complementary bases are added replacing T with U. • 4. As the RNA strand is synthesized it comes off of the template and the DNA strands rewind. • Once the RNA strand is finished, it folds into a shape that gives it its final function.

10. Translation • mRNA codes for functional proteins • 1. Ribosomes bind to the mRNA. • Each ribosome has 2 assembly sites amino acids. • Each protein is made up of a string of amino acids bound together. • 3 nucleotides of mRNA is called a codon • A codon codes for 1 amino acid. • There is also a start codon, which signals to the ribosomes and tRNA that translation starts here, and a stop codon. The stop codon doesn’t code for any amino acids. It is like a bump in the road that bumps the ribosomes off when translation is done.

11. 2. tRNA binds to an amino acid and takes it to the ribosome. • Each tRNA has one that binds to a specific amino acid. It can never bind to any other kind of amino acid. • The other end of the tRNA has an anti-codon. The anti-codon is complimentary to a specific codon on the mRNA. • So, the tRNA binds to its specific amino acid, then goes to the ribosome. It then enters the open assembly site and if the mRNA codon and the tRNA anti-codon match, then the amino acid is bound to its neighboring amino acid in the adjacent assembly site. (See figure 9.13)

12. 3. Amino acid elongation • As the amino acids are synthesized, the ribosome moves down the mRNA one codon at a time. This frees up one assembly spot in the ribosome for a new tRNA to bring a new amino acid. • Amino acid elongation occurs until the ribosomes have traveled down the length of the mRNA. Then the new amino acid chain is release and the ribosomes fall off of the mRNA. • 4. Protein folding • The newly formed chain of amino acids has many different charges on it due to the variety of chemical structures of the amino acids. Once the chain is released from the ribosome, it then folds into a functional shape based upon the charges and shapes of the amino acids. (Remember that negative repels negative, positive repels positive, and negative and positive attract.)