Microbial Genetics Lecture 1-2024 PDF

Summary

These lecture notes cover microbial genetics, including definitions of classical and molecular genetics. The document also details the structure of DNA, RNA, and various types of RNA. The notes also discuss the process of DNA replication.

Full Transcript

MICROBIAL GENETICS Dr. Amira El-Ganiny Professor of Microbiology & Immunology Lecture 1 Definitions Genetics: branch of biology that study Genes, Genetic variation and Heredity in living organisms  Classical genetics: old branch of genetics based only...

MICROBIAL GENETICS Dr. Amira El-Ganiny Professor of Microbiology & Immunology Lecture 1 Definitions Genetics: branch of biology that study Genes, Genetic variation and Heredity in living organisms  Classical genetics: old branch of genetics based only on visible results of reproductive acts  based on experiments of Mendel  Molecular genetics New field of genetics that studies structure & function of Nucleic acids at molecular level  Chromosomes structures, replication, gene expression Molecular GENETICS The nucleic acids  DNA and RNA are the carrier of genetic material in all living organisms. Nucleic acids have two important functions Ability to express genetic Ability to duplicate itself  traits through  transfer to progeny  by  transcription to mRNA chromosomal replication  translation into proteins. The Structure of DNA  DNA consists of nucleotides  3 parts  backbone of alternating phosphate & sugar (2-deoxyribose)  Nitrogenous base  Purines : adenine (A), guanine (G)  Pyrimidines: cytosine (C), thymine (T) A paired with T (2 hydrogen bonds) G paired with C (3 hydrogen bonds)  nucleotides are joined by  phospho- diester bond  DNA is double helix  2 chains of nucleotides coiled around each other  the 2 standards are complementary  The 2 chains are antiparallel (i.e., their sugar-phosphate backbones are oriented in opposite directions)  5' to 3' direction  3' to 5' direction  In prokaryotes  DNA is circular, super-coiled molecule associated with basic proteins (histone-like)  In eukaryotes  DNA is more organized  contain histone proteins & coiled into repeating units known as nucleosomes. RNA RNA is single strand  coil back on itself RNA composed of the sugar ribose  instead of 2- deoxyribose Uracil (U)  instead of thymine (T) There are 3 main types of RNA Ribosomal (rRNA) Transfer (tRNA) Messenger (mRNA) Types of RNA 1. Messenger RNA (mRNA)  Carry message  direct protein synthesis in ribosomes  mRNA  formed by transcription of DNA  Transcription depend on RNA polymerase  initiates RNA synthesis at promoter site on gene stop when termination codon reached. 2. Ribosomal RNA (rRNA) rRNAs are components of ribosomes They are made from large precursors  Enzymatically cleaved to  16s, 23s and 5s rRNA 3. Transfer RNA (tRNA) Carry amino acids  during protein synthesis tRNAs can distinguish different amino acids Each type of tRNA is covalently bound to one of the 20 aa. Each tRNA has a triplet of nucleotides called 'anticodon‘  binds to triplet nucleotides on mRNA, called codon during protein synthesis. When aa is attached to tRNA  tRNA is said to be charged. DNA REPLICATION Each strand of DNA serves as template for production of another complementary strand This pattern of replication is responsible for maintaining (conserving) proper sequence of bases on DNA molecule The pattern of DNA replication is described as 'semiconservative‘  produce 2 copies of DNA molecule  each copy contained one original strand and one new strand Replication starts at a point called 'origin of replication' by separation of the two strands  The replication origin  specific segment of DNA molecule consisting of about 245 bp. Replication fork  area of DNA molecule where strand separation occurs and the synthesis of new DNA takes place. A replicon  consists of origin of replication and DNA that is replicated from that origin  Bacterial chromosome has single replicon (one bubble).  Eukaryotes have multiple replicons (several bubbles exist) to efficiently replicate the relatively large molecules within a reasonable time Polymerization of nucleotides occur in 5' to 3' direction  for original strand Leading strand (3'  5‘) A challenge in DNA replication is  how to achieve 5‘ to 3' polymerization in the opposite direction from the template strand which is itself is from 5'-3' direction (lagging strand) This problem is solved by having different modes of polymerization for the two growing strands Leading strand Lagging strand Formed continuously Formed from 5‘3' end in small from the 5' to 3'end fragments  Okazaki fragments  linked together by ligase enzyme DNA replication DNA replication  carried out using DNA polymerases  cells in all organisms contain multiple highly specialized DNA polymerases Bacteria  5 DNA polymerases  I, II, III, IV, V  Yeast  8 DNA polymerases humans  at least 15 DNA polymerases The rate of DNA synthesis 750-1000 bp/second  in Prokaryotes  50-100 bp/second  in Eukaryotes  Polymerase III  main polymerase in DNA replication process  To initiate replication DNA polymerase require presence of primer  short strand of RNA to which growing polynucleotide chain is covalently attached  Polymerase I has exonuclease activity to remove mismatched nucleotide & adds correct nucleotide  repair process  Proofreading  removal of incorrect nucleotides immediately after they are added to growing DNA during replication process.  The proofreading function of DNA polymerase I improves fidelity of replication to one error in every 109 -1010 bp Steps of DNA replication in E. coli (1) DnaA protein binds to origin of replication (Ori C)  DNA replication starts by separation of the two strands. (2) Helicase(dnaB) bind replication fork to unwind the 2 DNA strands at same time topoisomerases (DNA gyrase) relieve the tension caused by unwinding process (3) Single-stranded DNA binding proteins (SSBs)  keep the single stranded region of the template DNA apart (4) Primase (dnaG)  synthesize small RNA molecule (~10 nucleotides)  act as primer for DNA synthesis (5) DNA polymerase III synthesizes complementary strand of DNA according to base-pairing rules The direction of DNA synthesis is from 5' to 3' end of the newly formed strand On one strand (leading strand), synthesis is continuous on the other (lagging strand) discontinuous synthesis (Okazaki fragments) are generated DNA synthesis is bi-directional  2 replication forks in opposite directions from origin of replication (6) DNA polymerase I  removes primer and fills gaps that result from RNA deletion. (7) DNA ligases join DNA fragments to form a complete DNA strand DNA replication is extraordinary complex; at least 30 proteins are required to replicate the E. coli chromosome.

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