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Microbial Biotechnology. Chapter 5. The Structure of Microbes. Prokaryotes Archaebacteria Includes halophiles, thermophiles, “extremophiles” Eubacteria On skin, soil, water, can be pathogenic. The Structure of Microbes. Characteristics of Prokaryotes Generally smaller than Eukaryotes

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The structure of microbes
The Structure of Microbes

  • Prokaryotes

    • Archaebacteria

      • Includes halophiles, thermophiles, “extremophiles”

    • Eubacteria

      • On skin, soil, water, can be pathogenic


The structure of microbes1
The Structure of Microbes

  • Characteristics of Prokaryotes

    • Generally smaller than Eukaryotes

    • No nucleus

    • Cell wall composed of peptidoglycan

    • Conjugation (transfer of DNA by cytoplasmic bridge)

    • Transduction (DNA is packaged in a virus and infects recipient bacterial cells)

    • 20 minute growth rate (binary fission


Yeast are important too
Yeast are Important Too!

  • Single celled eukaryote

  • Kingdom: Fungi

  • Over 1.5 million species

  • Source of antibiotics, blood cholesterol lowering drugs

  • Able to do post translational modifications

  • Grow anaerobic or aerobic

  • Examples: Pichia pastoris (grows to a higher density than most laboratory strains), has a no. of strong promoters, can be used in batch processes


Microorganisms as tools
Microorganisms as Tools

  • Microbial Enzymes

    • Taq (DNA polymerase), cellulases, proteases, amylases


Microorganisms as tools1
Microorganisms as Tools

  • Bacterial Transformation

    • The ability of bacteria to take in DNA from their surrounding environment

    • Bacteria must be made competent to take up DNA


Microorganisms as tools2
Microorganisms as Tools

  • Cloning and Expression Techniques

    • Fusion Proteins


Microorganisms as tools3
Microorganisms as Tools

  • Microbial Proteins as Reporters

    • Examples: the lux gene which produces luciferase

    • Used to develop a fluorescent bioassay to test for TB


Microorganisms as tools4
Microorganisms as Tools

  • Yeast Two-Hybrid System

    • Used to study protein interactions


Using microbes for a variety of everyday applications
Using Microbes for a Variety of Everyday Applications

  • Food Products

    • Rennin used to make curds (solid) and whey in production of cheese

    • Recombinant rennin is known as chymosin (first recombinant food product approved by FDA)


Using microbes for a variety of everyday applications1
Using Microbes for a Variety of Everyday Applications

  • Food Products

    • Energy production in bacteria

      • Aerobic or anaerobic


Using microbes for a variety of everyday applications2
Using Microbes for a Variety of Everyday Applications

  • Food Products

    • Fermentation

    • (anaerobic respiration)

    • Lactic acid fermentation

      • Used to make cheese, yogurt, etc.

    • Ethanol fermentation

      • Used to make beer and wine


Using microbes for a variety of everyday applications3
Using Microbes for a Variety of Everyday Applications

  • Therapeutic proteins

    • Recombinant insulin in bacteria


Using microbes for a variety of everyday applications4
Using Microbes for a Variety of Everyday Applications

  • Field Applications of Recombinant Microorganisms

    • Ice-minus bacteria (remove ice protein producing genes from P. syringae)

    • P. fluorescens containing the gene that codes for the bacterial toxin from Bacillus thuringiensis (kills insects) Bt toxin!


Using microbes for a variety of everyday applications5
Using Microbes for a Variety of Everyday Applications

  • Using Microbes Against Other Microbes

    • Antibiotics

    • Act in a few key ways

      • Prevent replication

      • Kill directly

      • Damage cell wall or prevent its synthesis


Vaccines
Vaccines

  • First was a vaccine against smallpox (cowpox provides immunity)

    • DPT-diphtheria, pertussis, and tetanus

    • MMR –measles, mumps, and rubella

    • OPV- oral polio vaccine (Sabin)


Vaccines1
Vaccines

  • A Primer on Antibodies

    • Antigen- foreign substances that stimulate an immune response

    • Types of leukocytes or white blood cells

      • B-lymphocytes: antibody-mediated immunity

      • T-lymphocytes: cellular immunity

      • Macrophages: “cell eating” (phagocytosis)


Vaccines

  • Antigens stimulate antibody production in the immune system

Light chain

Heavy chain

IgA – first line of defense

IgG and IgM – activates macrophages


Vaccines

  • Mechanism of Antibody Action


Vaccines

  • How are vaccines made?

    • They can be part of a pathogen (e.g. a toxin) or whole organism that is dead or alive but attenuated (doesn’t cause disease)

      • Subunit (toxin) or another part of the pathogen

      • Attenuated (doesn’t cause disease)

      • Inactivated (killed)

  • What about flu vaccines (why do we have to get a shot every year?)


Bacterial and Viral Targets for Vaccines

  • Immune Therapy

    • Used to enhance natural immune responses

    • Ex: Interferon is a protein that triggers production of protective proteins (or antibodies) that inhibit viral replication.



Microbial genomes
Microbial Genomes

  • Microbial Genome Program (MGP) –the goal is to sequence the entire genomes of microorganisms that have potential applications in environmental biology, research, industry, and health

  • Sequencing Strategies


Microbial genomes1
Microbial Genomes

  • Why study viral genomes?

    • Decipher genes and their products so that agents that block attachment, block replication can be made


Microbial diagnostics
Microbial Diagnostics

  • Using Molecular Techniques to Identify Bacteria

    • RFLP

    • PCR and Real time PCR

    • Sequencing


Microbial diagnostics1
Microbial Diagnostics

  • Microarrays for tracking contagious disease

    • PulseNet used to identify outbreaks


Microbial diagnostics2
Microbial Diagnostics

  • Combating Bioterrorism

    • The use of biological materials as weapons to harm humans or animals and plants we depend on for food

    • Examples in History

      • Throwing plague infected dead bodies over the walls of their enemies


Microbial diagnostics3
Microbial Diagnostics

  • Using Biotech Against Bioweapons

    • Postal service x-raying packages

    • Antibody tests in the field

    • PCR tests in the field

    • Protein Microarrays for detecting bioweapon pathogens


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