Food Microbio Exam 2 Flash Cards

Ch 5: Metabolism

• Study of chemical reactions by which microbes grow and reproduce

• catabolism/catabolic or degradative reactions: chemical reactions that release energy by breakdown of nutrients, provide energy and building blocks for anabolism

• anabolism/anabolic or biosynthetic reactions: use energy and building blocks to build large molecules and for other cellular functions

• Most microorganisms are chemoorganotrophs (use organic molecules) or chemolithotrophs (use inorganic molecules).

Metabolism

• Buildup and breakdown of chemicals in a cell via chemical reactions that generate energy and substances that sustain life

• All biological systems convert energy into aTP which is used to power biosynthetic reactions

• A metabolic pathway is a sequence of enzymatically catalyzed chemical reactions in a cell

• Metabolic pathways are determined by enzymes

• Enzymes are encoded by genes

Gibb’s free energy change

• The amount of available energy liberated or used up in a reaction

• If change in G is negative; exergonic; reactions that release energy

  • Eg. oxidation of glucose to CO2 and H2O

• If change in G is positive; endergonic; reactions that require energy

  • Eg. protein synthesis

• Free energy change of breakdown or hydrolysis is usually reported as kcal/mol

ATP

• High energy carrier molecule

• All biological systems convert energy into ATP which is used to power biosynthetic reactions

• Energy released from chemical reactions captured in energy rich chemical bonds in ATP, AMP, UTP

• Unstable anhydride bonds between the phosphate groups formed by: ADP + H3PO4 → ATP + H2O; change in G= +7.3 kcal/mol

Phosphorylation reactions

• Substrate level phosphorylation

• Oxidative phosphorylation

• Photophosphorylation

Substrate Level Phosphorylation

• Energy from the transfer of a high energy PO4 to ADP; generates ATP

Oxidative Phosphorylation

• Energy released from transfer of electrons (oxidation) of one compound to another (reduction) is used to generate ATP in the electron transport chain

• Occurs in plasma membrane

Photophosphorylation

• Light causes chlorophyll to give up electrons. Energy released from transfer of electrons (oxidation) of chlorophyll

Pyridine Nucleotides

• Nicotinamide adenine dinucleotide (NAD): a coenzyme that is also involved in energy reactions

• An electron carrier: reduced state (NADH) or oxidized state (NAD+)

  • NAD+ + 2e- + 2H+ → NADH+ + H+

• NADH is a strong reducing agent that participates in many enzyme-catalyzed oxidation-reduction reactions necessary in cell metabolism

Oxidation-Reduction

• Oxidation: removal of electrons

• Reduction: gain of electrons

• Redox reaction: an oxidation reaction paired with a reduction reaction

Biological Oxidation

• In biological systems, electrons are often associated with hydrogen atoms; biological oxidations are often dehydrogenations

Enzymes

• Biological catalysts

  • Specific for a chemical reaction; not used up in that reaction

• Will only function under their optimum temperature and pH ranges

• Apoenzyme: Protein

• Cofactor: nonprotein component

  • Coenzyme: organic cofactor

• Holoenzyme: apoenzyme plus cofactor (whole active enzyme)

Important coenzymes

• Carriers of functional groups, specific atoms, electrons

• Nonspecific, recognized by many enzymes that compete for their availability

• Not all enzymes require coenzymes, many do

• Participate in redox rxns, transfer of chemical groups, molecular rearrangement reactions

  • NAD+

  • NADP+

  • FAD

  • Coenzyme A

Enzyme classification

• Oxidoreductase: oxidation-reduction

• Transferase: transfer functional groups

• Hydrolase: hydrolysis

• Lyase: removal of atoms without hydrolysis

• Isomerase: rearrangement of atoms

• Ligase: joining of molecules, uses ATP

Factors affecting enzyme activity

• pH

• Temperature

• Substrate concentration

• Inhibitor

Inhibitors

• Competitive

• Non-competitive

• Feedback

Carbohydrate catabolism

• Glycolysis

• Krebs cycle

• Electron transport chain

Glycolysis

• The oxidation of glucose to pyruvic acid produces ATP and NADH

Overall glycolysis reaction: Glucose + 2 ATP + 2 ADP + 2 PO4- + 2 NAD+ → 2 pyruvic acid + 4 ATP + 2 NADH + 2 H+

Alternative to Glycolysis

• Pentose phosphate pathway

  • Uses pentoses and NADPH

  • Operates with glycolysis

• Entner doudoroff pathway

  • Produces NADPH and ATP

  • Does not involve glycolysis

  • Pseduomonas, Rhizobium, Agrobacterium

Cellular respiration

• Oxidation of molecules liberates electrons for an electron transport chain

• ATP is generated by oxidative phosphorylation

Krebs/TCA Cycle

• Oxidation of acetyl CoA to produce NADH, FADH2 and GTP/ATP

Electron Transport Chain

• A series of carrier molecules that are in turn oxidized and reduced as electrons are passed down the chain

• Energy released can be used to produce ATP by chemiosmosis

• 3 classes of carriers

  • Flavoproteins: protein, contain flavin (coenzyme from riboflavin), eg. FMN

  • Cytochromes: protein, contain heme group, can exist in reduced (Fe 2+) and oxidized (Fe 3+) form

  • Ubiquinones or coenzyme Q: non protein

Summary of respiration

• Aerobic: final electron acceptor in the ETC is molecular oxygen

• Anaerobic: final electron acceptor in ETC is not O2. Yields less energy than aerobic respiration because only part of the Krebs cycle operates under anaerobic conditions

Fermentation

• Scientific

  • Releases energy from oxidation of organic molecules

  • Does not require oxygen

  • Does not use the Krebs Cycle or ETC

  • Uses an organic molecule as the final electron acceptor

• General

  • Any spoilage of food by microorganisms

  • Any process that produces alcoholic beverages or acidic dairy products

  • Any large scale microbial process occuring with or without air

• Alcohol fermentation

  • Produces ethanol and CO2

• Lactic acid fermentation

  • Produces lactic acid

    • Homolactic: produces lactic acid only

    • hetero lactic: produces lactic acid and other compounds

Microbial Nutrition and Growth

Requirements for growth

• Physical

  • Temperature

  • pH

  • Osmotic pressure

• Chemical

  • Carbon

  • Nitrogen, sulfur, and phosphorus

  • Oxygen

  • Trace elements

  • Organic growth factor

• Carbon

  • Structural organic molecules, energy source

  • Chemoheterotrophs use organic carbon sources

  • Autotrophs use CO2

• Nitrogen

  • In amino acids and proteins

  • Most bacteria decompose proteins

  • Some bacteria use NH4 or NO3

  • A few bacteria use N2 in Nitrogen fixation

• Sulfur

  • In amino acids, thiamine and biotin

  • Most bacteria decompose proteins

  • Some bacteria use SO4 or H2S

• Phosphorus

  • In DNA, RNA, ATP and membranes

  • PO4 is a source of phosphorus

• Trace elements

  • Inorganic required in small amounts

  • Usually as enzyme cofactors

• Organic growth factors

  • Organic compounds obtained from the environment

  • Vitamins, amino acids, purines, and pyrimidines

  • Fastidious microbes: cannot grow in minimal media

• Culture media

  • Culture medium: nutrients prepared for growth

  • Sterile: no living microbes

  • Inoculum: introduction of microbes into medium

  • Culture: microbes growing in/on culture medium

• Agar

  • Complex polysaccharide

  • Use as solidifying agent for culture media in petri plates, slants, and stabs

  • Generally not metabolized by microbes

  • Liquifies at 100C

  • Solidifies around 40C

• Culture media

  • Chemically defined media: exact chemical composition is known

    • Usually minimal media, fastidious microbes cannot grow

  • Complex media: extracts and digests of yeasts, meat, or plants

    • Nutrient broth

    • Nutrient agar

• Selective media

  • Addition of selective ingredients eg. antibiotics or dyes

  • Suppress unwanted microbes and encourage desired microbes

• Differential media

  • Addition of specific ingredients eg. certain carbs and pH indicator dyes

  • Make it easy to distinguish colonies of different microbes

Living Cells

• To grow parasitic microbes

• Can be mammalian cells, embryos, even live animals

Enrichment culture

• Encourages growth of desired microbe or microbes that have been injured

• Can be done in the step prior to plating on selective medium

Aseptic techniques

• To transfer pure cultures without allowing contamination to occur from other microbes

• Use of sterile equipment materials

Pure cultures

• A pure culture contains only one species or strain

• A colony is a population of cells arising from a single cell or spore or from a group of attached cells

• A colony is often called a colony forming unit (CFU)

• The streak plate method is used to isolate pure cultures

Preserving bacterial cultures

• Agar slants/stabs: 3-5C

• Deep freezing: -50 to -95C

• Liquid nitrogen: -190C

• Lyophilization (freeze dry): -54 to -72C and dehydrated in vacuum

• ATCC: American type culture collection: culture repository

pH

• Most bacteria grow between 6.5 and 7.5

• Molds and yeasts grow between pH 5 and 6

• Acidophiles grow in acidic environments

• Sulfolobus, thermoplasma, thiobacillus: obligate acidophiles

Atmospheric conditions (Oxygen)

Anaerobic culture methods

• Reducing media

  • Contain chemicals (thioglycolate or oxyrase) that combine O2

  • Heated to drive off O2

• Anaerobic jars, chambers, glove boxes

Temperature

Hydrostatic pressure

• Barophiles are microbes that grow best at high pressure

• Deep sea bacteria

Osmotic pressure

• Hypertonic environments, or an increase in salt or sugar, causes plasmolysis

• Extreme or obligate halophiles require high osmotic pressure

• Facultative halophiles tolerate high osmotic pressure

• Osmohpiles are tolerant of high pressure

Reproduction in prokaryotes

• Binary fission

• Budding

• Conidiospores (actinomycetes)

• Fragmentation of filaments

Generation time

• = t/3.3log (b/B)

  • t= time between initial and end

  • b= end count

  • B= initial count

  • 3.3= log2 to log10 conversion factor

Continuous culture

• Growing a culture continuously

• A chemostat or bioreactor is used

• Keep the inflow of nutrients outflow of waste constant

• Industrial fermentations are continuous cultures