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“微生物学”考试时间地 点 时间:2000年1月9日上午8:00-10:00 地点:四教4206 Detailed phylogenetic tree of the Archaea based on 16S ribosomal RNA sequence Comparisons Archaeal Membranes and Cell Wall Archaea lack fatty acids, instead have hydrocarbon moieties bonded to glycerol by ether (instead of ester) linkages Glycerol diethers and diglycerol tetraethers are the major classes of lipids present in Archaea Archaea do not contain muramic acid and D-amino acids, as in Bacteria A pseudopeptidoglycan is found in some archaea, it consists of two amino sugars: N-acetylglucosamine and Nacetyltalosaminuronic acid, with only L-amino acids linkages Some contain a thick wall consists only polysaccharide Some contain cell walls made of glycoprotein Some lack carbohydrate in their cell walls and have walls consisting of only protein. Chapter 20 Prokaryotic Diversity: Archaea Extremely Halophilic Archaea Methane-Producing Archaea: Methanogenes Hyperthermophilic Archaea Thermoplasma: A Cell-Wall-Less Archaean Limits of Microbial Existence: Temperature Archaea: Earliest Life Forms? Extremely Halophilic Archaea: inhabitants of highly saline environments such as solar salt evaporation ponds and natural salt lakes Hypersaline habitats: Seawater evaporating ponds: the red-purple Great Salt Lake in Utah Color is due to bacterioruberins and bacteriorhodopsin of halobacterium Environments for extremely halophile Solar salt evaporation ponds Natural salt lakes Artificial saline habitats (surfaces of heavily salted food such as certain fish and meats) Require at least 1.5 M (9%) NaCl for growth Most species require 2-4 M (12-23%) NaCl for growth Some can grow at pH of 10-12 No harmful to human and animals Physiology of Extremely Halophilic Archaea All are chemoorganotrophs Most are obligate aerobes All require large amount of sodium for growth All stain gram negatively, binary fission growth Most are nonmotile Halobacterium and Halococcus contain large plasmids Peptidoglycan replaced by glycoportein in the cell wall Cellular components exposed to the external environment require high Na+ for stability Cellular internal components require high K+ for stability Na+ stabilize the cell walls. Bacteriorhodopsin and Light-mediated ATP Synthesis Bacteriorhodopsin Methane-Producing Archaea: Methanogens Methane formation occurs under strictly anoxic conditions. CO2-type substrates (CO2, HCOO- and CO) can be used as carbon sources. Methyl substrates (CH3OH, CH3NH2+, (CH3)2NH+, (CH3)3NH+, CH3SH, (CH3)2S) are methanogenic carbon sources. Acetotrophic substrates such as acetate can also be used to produce methane. Three classes of methanogenic substrates are known and all release free energy suitable for ATP synthesis Diversity and Physiology of Methanogenic Archaea 16S ribosomal RNA sequence analyses classify methanogen into seven major groups All methanogens use NH4+ as a nitrogen source A few species can fix molecular nitrogen Nickel is a trace metal required by all methanogens, it is a component of coenzyme Factor430 Iron and Cobalt are also important for methanogens. Pictures on the left: morphological diversity of methanogens Diversity and Physiology of Methanogenic Archaea Picture on the left are hyperthermophilic and thermophilic methanogens Methanococcus jannaschii (85oC optimal) Methanococcus igneus (88oC optimal) Methanothermus fervidus (85oC optimal) Methanothrix thermophila 60oC optimal) Picture on the right: thin section of methanogenic Archaea: Methanobrevibacter ruminantium Methanosarcina barkeri Unique Methanogenic Coenzymes Methanofuran (MF): a low-molecular-weight coenzyme that interacts in the first step of methanogenesis from CO2. Methanopterin (reduced form tetrahydro-methanopterin or MF): a methanogenetic coenzyme containing a substituted pterin (蝶呤) ring, a C1 carrier during the reduction of CO2 to CH4. Coenzyme M: involved in the final step in methane formation, is the carrier of the methyl group that is reduced to methane by the F430-methyl reductase enzyme complex in the final step of methanogenesis. Coenzyme F430: a yellow, soluble, nickel-containing tetrapyrrole that plays an intimate role in the terminal step of methanogenesis as part of the methyl reductase system. Unique Methanogenic Coenzymes Coenzymes involved in redox reactions Coenzyme F420: an electron donor in methanogenesis. 7-mercaptoheptanoylthreonine phosphate (HSHTP): an electron donor in methanogenesis, is the final unique coenzyme of the methanogens to be considered. Coenzymes unique to methanogenic Archaea Coenzymes Unique to Methanogenic Archaea The oxidized form of F420 absorbs light at 420 nm and fluresces blue-green. On reduction, the coenzyme becomes colorless. The fluorescence of F420 is a useful tool for preliminary identification of an organism as a methanogen Autofluorescence of the methanogen Methanosarcina barkeri due to the presence of the unique electron carrier F420. Pathway of methanogenesis from CO2 Autotrophy in Methanogens C1-carrying corrinoidcontaining enzyme How autotrophic methanogens combine aspects of biosynthesis and bioenergetics. Note how half of the acetyl-CoA molecule produced comes from reactions leading to methanogenesis. Methanogenesis from methyl compounds and acetate Utilization of reactions of the acetyl-CoA pathway during growth on methanol (a) acetate (b) Energetic of Methanogenesis ATP synthesis linked to a proton motive force established during the terminal step of methanogenesis Hyperthermophilic Archaea Temperature Optima above 80oC Most isolated from geothermally heated soils or waters containing sulfur an sulfides Most are obligate anaerobes Many grow chemolithotrophically, with H2 as energy source Hyperthermophilic from Volcanic Habitats Acidophilic Hyperthermophilic Archaea The first such organism discovered, Sulfolobus, grows in sulfur-rich hot acid springs at temperature up to 90oC and at pH values of 1-5. Acidianus, a facultative aerobe resembling Sulfolobus is also present in acidic solfataric springs, it can also grow anaerobically. Acidianus infernus Sulfolobus acidocaldarius Hyperthermophilic from Volcanic Habitats Acidophilic Hyperthermophilic Archaea Spherical, obligately anaerobic, S0-respiring organism. Grows best at neutral pH and 80-90oC Desulfurococcus saccharovorans Hyperthermophilic from Volcanic Habitats Acidophilic Hyperthermophilic Archaea Thermoproteus and Thermofilum inhabit neutral or slightly acidic hot springs, are highly variable in length, ranging from 1-80 microns. Both are strict anaerobes that carry out a S0-based anaerobic respiration. Most can grow chemolithotrophically. Thermoproteus neutrophilus Thermofilum librum Thermofilum librum Hyperthermophilic from Submarine Volcanic Areas Boiling points increase with water depth. Pyrodium has a growth optimum of 105oC, has higher GC(62%). Cells are irregularly disc- and dish-shaped, grow in culture as a moldlike layer on sulfur crystals suspended in the medium. Strict anaerobe that grows chemolithotrophically at neutral pH on H2 with S0 as electron acceptor. Growth occur between 82-110oC. Pyrodium occultum (optima 105oC) Hyperthermophilic from Submarine Volcanic Areas Pyrobaculum is capable of both aerobic respiration and denitrification (NO3N2). Organic or inorganic substrates can be used as electron donors Maxima T=103oC H2, as well as various complex nutrients but not sugars support its growth. Elemental So is not used by this organism, even inhibits its growth. Pyrobaculum aerophilum (optima 100oC) Hyperthermophilic from Submarine Volcanic Areas Thermococcus, a spherical hyperthermophilic archaean indigenous to anoxic submarine thermal waters in various location worldwide. Contains a tuft of polar flagella, highly motile. Obligately anaerobic chemoorganotroph that grows on proteins and other complex organic mixtures (including some sugars) with Hyperthermococcus celer Dividing cells of S0 as electron acceptor. Pyrococcus furiosus Optima T=88oC Pyrococcus grows at between 70106oC with an optimum of 100 oC. Metabolic requirement similar to Hyperthermococcus. Hyperthermophilic from Submarine Volcanic Areas Staphylothermus consists of spherical cells about 1 micron in diameter that form aggregates of up to 100 cells. Strictly anaerobic hyperthermophile growing optimally at 92oC. Capable of growth between 65 and 98oC. S0 is required for growth, yet oxidation of complex organic compounds is not tightly coupled to S0 reduction. Staphylothermus marinus Hyperthermophilic from Submarine Volcanic Areas Most Archaea use S0 as an electron acceptor for anoxic growth, most are unable to use sulfate as an electron acceptor. Archaeoglobus, is a true sulfate-reducing hyperthermophile. Grow at between 64 and 92oC with T optima=83oC Share some metabolic features with methanogens. Methanopyrus kandleri Archaeoglobus lithotrophicus Methanopyrus: gram-positive rod-shaped methanogen grown above 100oC. The most ancient hyperthermophile Share phenotypical properties with both the hyperthermophiles and methanogens. Hyperthermophilic from Submarine Volcanic Areas Aquifex and Thermotoga are not Archaea but hyperthermophilic bacteria that otherwise strongly resemble hyperthermophilic Archaea. Thermotoga maritima (80oC) Chemoorganotrophic and anaerobic Aquifex pyrophilus (85oC) Obligate chemolithotrophic, microaerobically or anaerobically growth with only H2, S0 or S2O3- as electron donor and O2 or NO3- as electron acceptor. Thermoplasma: A Cell-Wall-Less Archaea Thermoplasma acidophilum an acidophilic, thermophilic mycoplasma-like archaea Thermoplasma volcanium has been isolated from Solfatara fields throughout the world. Thermoplasma volcanium shadowed preparation Thermoplasma acidophilum is a cell-wall-less prokaryote resembling the mycoplasmas. Acidophilic, aerobic chemoorganotroph, thermophilic Archaea (pH=2 and To=55oC). All strains of Thermoplasma have been isolated from self-heating coal refuse piles. Thermoplasma: A Cell-Wall -Less Archaea Self-heating coal refuse pile habitat of Thermoplasma Thermoplasma has evolved a cell membrane of chemically unique structure. It contains lipopolysaccharide consisting of a tetraether lipid with mannose and glucose units. The membrane also contains glycoproteins but not sterol, the overall structure render the thermoplasma membrane stable to hot acid conditions Limits of Microbial Existence: Temperature Pyrodictium occultum (optima 105oC, maxima 110oC) Laboratory experiments on the heat stability of biomolecules suggest that living processes could be maintained at temperature as high as 140-150oC. Structure of the tetraether lipoglycan of Thermoplasma acidophilum Archaea: Earliest Life Forms? Early geochemical conditions: High temperature High salt Low pH Strict anoxic conditions Only Archaea can stand such environmental extrems. Do you agree with the argument: Archaea are the Earliest Life Forms