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Uptake of nutrients Cells require nutrients to grow and survive Cells possess numerous transport systems Must deal with specificity, transport against conc. gradient and transport across selective membrane Passive vs. facilitated diffusion Diffusion = movement from region of high conc. to region of low conc. Some molecules can pass through the membrane (e.g. H2O, O2, CO2) passive diffusion Passive vs. facilitated diffusion Many molecules require a carrier protein (permease) to cross the membrane At high conc. permeases are all occupied (plateau in transport rate) Passive vs. facilitated diffusion Permeases are selective and only transport closely related molecules Permeases span the entire membrane Model of facilitated diffusion Binding of substrate to permease may induce conformational change, releasing molecule in the cell interior Permease would then change back and bind another molecule Passive and facilitated diffusion Both require that nutrient be in higher conc. outside cell Prokaryotes usually grow in nutrient-poor environments Diffusion more common in eukaryotes Active transport and group translocation Prokaryotes expend energy to transport nutrient against a conc. gradient Can use active transport or group translocation Active transport ATP-binding cassette (ABC) transporters Consist of 2 pore-forming membrane-spanning domains Hydrolysis of ATP drives transport Active transport ATP-binding cassette (ABC) transporters Often work with periplasmic (or external) substrate-binding protein Transport through outer membrane is via porins, or outer membrane receptors/transporters Active transport Proton gradients can drive active transport Symport: substrate and proton both transported into the cell Antiport: Na+ transported out as proton enters cell Uptake of Na+ drives uptake of nutrient = symport Group translocation Molecule is chemically altered as it is being transported into the cell Requires energy Phosphoenolpyruvate: sugar phosphotransferase system (PTS) Example of group translocation Transports various sugars Consists of two enzymes and a low molecular weight protein High energy phosphate transferred from PEP via enzymes to sugar molecule PEP + sugar (outside) pyruvate + sugar-P (inside) Iron uptake Iron is required by almost all prokaryotes Siderophores are low molecular weight iron-binding molecules secreted by microorganisms Either iron or iron-siderophore complex is transported into cell by ABC transporters Iron uptake Periplasmic iron-binding proteins and inner membrane transporters required in gramnegative bacteria Nutrient requirements All cells require specific elements to survive Macroelements Carbon, Oxygen, Hydrogen, Nitrogen, Sulfur, Phosphate, Potassium, Calcium, Magnesium and Iron Nutrient requirements All cells require specific elements to survive Macroelements Carbon, Oxygen, Hydrogen, Nitrogen, Sulfur, Phosphate, Are components of carbohydrates, proteins, lipids and nucleic acids Nutrient requirements All cells require specific elements to survive Macroelements Potassium, Calcium, Magnesium and Iron Exist as cations in the cell Play various roles (e.g. required for enzyme activity) Nutrient requirements All cells require specific elements to survive Trace elements Manganese, Zinc, Cobalt, Molybdenum, Nickel and Copper Contribute to enzyme activity (cofactors) Carbon requirement Can be satisfied by organic molecules (also contribute oxygen and hydrogen) Organic molecules can also serve as a source of electrons (energy) Heterotrophs use organic molecules as their carbon source Carbon requirement Can be satisfied by CO2 (oxidized form of carbon) Reduction of CO2 is energy expensive process Autotrophs use CO2 as their sole carbon source Energy source Can either be from light or the oxidation of organic or inorganic molecules Phototrophs obtain energy from light Chemotrophs obtain energy from oxidations Electron source Lithotrophs (rock eaters) obtain electrons from inorganic molecules Organotrophs obtain electrons from organic molecules Nitrogen requirement Used for the synthesis of various organic compounds Many microorganisms use amino acids and ammonia Most phototrophs reduce nitrate to ammonia and then use ammonia Some bacteria can reduce and assimilate atmospheric nitrogen Phosphorus requirement Required for the synthesis of various cell components like phospholipids and ATP Almost all microorganisms can use inorganic phosphate as source E. coli uses both inorganic and organic sources Sulfur requirement Required for the synthesis of cysteine, methionine and some carbohydrates and vitamins Most microorganisms use sulfate Some require cysteine (reduced form of sulfur) Growth factors Organic compounds that are required and cannot be synthesized by the organisms Amino acids Purines and pyrimidines Vitamins Vitamins Small organic molecules Often make up part or all of an enzyme cofactor Many vitamins are commercially produce using microorganisms Defined vs. complex culture media A defined or synthetic medium contains a known amount of specific chemical components Defined vs. complex culture media A complex medium contains some ingredients of unknown chemical composition Useful for growing a variety of microorganisms or when growth requirements of microorganisms are unknown Agar Most commonly used solidifying agent Sulfated polymer of sugars Extracted from red algae Selective vs. differential media Selective media favor the growth of particular microorganisms Differential media allow for the differentiation between different bacteria based on biological characteristics Isolation of pure cultures Required for the characterization of individual species Streak plate method Spread plate method Pour plate method (See figures in textbook)