MCB 100 Introductory Microbiology 2019 Microbial Genetics Chapter

MCB 100 Introductory Microbiology 2019 Microbial Genetics Chapter 7, 8 (pgs. 236 -246) The Central Dogma

MCB 100 Introductory Microbiology March 6, 2019 Chapter 7 Microbial Genetics Regulation of the E. coli Lac Operon

Fig. CO 07 DNA Double Helix

The “Semi-Conservative" Mechanism of DNA Replication is the duplication of a DNA molecule. Since DNA is composed of two strands that have a complementary sequence, each strand can act as a template for directing the synthesis of the other strand. The enzyme that makes new DNA is called DNA polymerase III. Replication is a bidirectional process. "Semi-conservative replication" means that the new double stranded DNA molecules consist of one strand that is from the old DNA molecule and one strand that is from the new DNA molecule.

Double stranded DNA

DNA Replication is a Complicated Process Materials Necessary for DNA Replication - Nucleotides: d. NTPs (deoxyribonucleotidetriphosphates) d. ATP, d. GTP, d. CTP and d. TTP - Template: A preexisting strand of DNA that determines the sequence of nucleotides in the new strand. - Primer: A short piece of DNA or RNA that binds to the template and has a free 3’ end. - Enzymes: . Helicase – separates the strands of old DNA. single-strand binding protein – keeps the two strands apart. primase – makes the short RNA primers. DNA polymerase III – adds nucleotides to the end of the primer. DNA polymerase I – removes primers, fills in gap. DNA ligase – joins pieces of DNA gyrase – controls super twisting of DNA topoisomerase – controls super twisting of DNA - Origin of replication – a sequence of DNA where the enzymes of replication bind and start the process of duplicating the DNA

Bacterial chromosome packaged in nucleoid

Before a cell divides, the genetic material must be completely copied so each daughter cell can get a chromosome. In Escherichia coli, at the optimal growth temperature, it takes about 100 minutes for a replication fork (a DNA polymerase complex) to go all of the way around the chromosome. At the optimal growth temperature, in a rich medium, the generation time for Escherichia coli, is about 30 minutes. How is this possible?

The E. coli chromosome is about 4 million base pairs long. Each replication fork copies about 2 million base pairs on both strands of the DNA. A replication fork moves at a rate of about 20, 000 base pairs per minute. Image from: www. expertsmind. com

When E. coli cells are growing at their fastest rate, new rounds of replication begin before the first round is completed.

Replication of DNA Which ONE of the following statements about the replication of DNA is FALSE? A. The energy required for synthesis of DNA comes from the hydrolysis of high-energy phosphoanhydride bonds. B. In replication of DNA new nucleotides are attached to the growing chain at the 3' end. C. Before a new strand of DNA can be made, the old strands have to be separated. D. The old strand of DNA acts as a template for the synthesis of a new strand which is "complimentary and antiparallel" to the old strand. E. The enzyme that is responsible for making new DNA is called the ribosome.

Replication of DNA Which ONE of the following statements about the replication of DNA is FALSE? A. The energy required for synthesis of DNA comes from the hydrolysis of high-energy phosphoanhydride bonds. B. In replication of DNA new nucleotides are attached to the growing chain at the 3' end. C. Before a new strand of DNA can be made, the old strands have to be separated. D. The old strand of DNA acts as a template for the synthesis of a new strand which is "complimentary and antiparallel" to the old strand. E. The enzyme that is responsible for making new DNA is called the ribosome.

Transcription and Translation

Transcription – the synthesis of RNA When RNA is synthesized, just one strand of the DNA is used as the template. The two strands of DNA are called "sense" and "antisense". The sense strand of the DNA has the same sequence as the m. RNA, it is not used as the template for RNA synthesis. The sequence of the antisense strand is complementary to the m. RNA, it is the template. Typical bacterial m. RNA molecules are 1000 – 3000 nucleotides long. The transcription bubble, where the strands are separated, is about 20 base pairs of DNA long. It moves down the DNA double helix so the RNA polymerase can read the whole gene and make a long m. RNA molecule.

The 3 Major Types of RNA m. RNA r. RNA t. RNA m. RNA - Messenger RNA, or m. RNA, is a copy of the information in a gene that is needed to direct the synthesis of a protein. The m. RNA consists of a sequence of codons, each codon specifies the incorporation of one particular amino acid in a growing protein chain. If a protein is 300 amino acids long, the m. RNA encoding that protein must be 300 codons long. Since each codon is 3 bases long, the m. RNA for a protein that has 300 amino acids must be at least 900 bases long. Messenger RNA is a single stranded molecule that is quickly degraded. In E. coli a typical m. RNA molecule lasts for only 1 - 2 minutes.

Transcription - Synthesis of RNA Choose the ONE statement about transcription as it occurs in Bacterial cells that is FALSE. A. A substance that inhibits bacterial RNA polymerase can kill a bacterial cell. B. The site where RNA synthesis begins is called a promoter. C. The strand of the DNA that is used as the template for the synthesis of a m. RNA molecule is called the "antisense" strand. D. The enzyme that makes m. RNA in bacteria is called an RNA-dependent DNA polymerase. E. The template used for the synthesis of RNA in a bacterial cell is DNA.

Transcription - Synthesis of RNA Choose the ONE statement about transcription as it occurs in Bacterial cells that is FALSE. A. A substance that inhibits bacterial RNA polymerase can kill a bacterial cell. B. The site where RNA synthesis begins is called a promoter. C. The strand of the DNA that is used as the template for the synthesis of a m. RNA molecule is called the "antisense" strand. D. The enzyme that makes m. RNA in bacteria is called an RNA-dependent DNA polymerase. The enzyme that makes m. RNA in all types of cells is a DNA-dependent RNA polymerase. E. The template used for the synthesis of RNA in a bacterial cell is DNA.

The 3 Major Types of RNA (continued) r. RNA - Ribosomal RNA, or r. RNA, is a structural part of the ribosome. The ribosome is basically a complex enzyme that synthesizes protein. The r. RNA molecules help to organize the 50 or so proteins that make up the ribosome and may be directly involved in the catalysis of reactions required for protein synthesis. RNA is a single stranded molecule but it can have sections of double-stranded structure if it has a sequence that allows it to fold back on itself in a hairpin-like structure. Ribosomal RNAs have a lot of regions of double stranded structure. This, and the fact that it is covered with ribosomal proteins make r. RNA much more stable than m. RNA.

The 3 Major Types of RNA (continued) t. RNA - Transfer RNA brings the correct amino acid into the ribosome and positions it in the place where it needs to be so the protein chain can grow. Transfer RNA has a "clover leaf"-like structure formed by hairpin formations. One of the loops contains the "anticodon" a segment of three nucleotides which is the complement to a specific codon. For more information about the process of translation and the role of m. RNA, r. RNA and t. RNA in protein synthesis see pages 212 – 219 of the text. There are 64 codons in the genetic code. Each codon is 3 bases long and there are 4 possible bases, 43 = 64. There is a specific t. RNA for each type of codon. The anticodon loop of the t. RNA binds to a complementary codon in the m. RNA. A specific amino acid is attached to the acceptor arm. The D-loop and Ty. C loop bind specific charging enzymes that attach the amino acid.

See Fig. 7. 14 Codon to anticodon binding between a m. RNA and a t. RNA