OCN621: Biological Oceanography- Bioenergetics-III

Transcription

1 OCN621: Biological Oceanography- Bioenergetics-III Guangyi Wang POST 103B dc 1 /dt = growth Standing Stock C 1 Overview 1. Energy Currency ATP 1) Active transport & ATP 2) Mechanical work & ATP 3) Biosynthesis i & ATP 2. Application of Bioenergetics to Biological Oceanography 1) Measurement of standing stocks (methods) 2) Measurement of rate processes 3) Relationship among cell constituents 4) Energetic efficiency i 5) Energy implications of adaptations 3. Assignment question 1) Link bioenergetics to physical and biogeochemical oceanographic processes???? 1

2 Biological Utilization of Chemical Energy 1. Energy Currency ATP - Economic analogy for the transformation of energy in the cell - need for a "medium of exchange". Most biochemical reaction series requires elaborate cell machinery and organization, and many specific enzymes. It is not efficient, and not possible, for enzyme complexes to handle all possible combinations of substrates, intermediates, and sources of energy. METABOLIC processes (e.g, respiration) "oxidize" organic molecules, capturing some of their energy in a single molecule which is recognized as the "energy donor" or "medium of exchange" in all subsequent reactions. This molecule, ATP, is special because it carries a fixed amount of energy in an easily released form high energy phosphate bonds. 2. Active Transport: work of moving molecules &ions against concentration gradients According to the 2nd Law of Thermodynamics - everything in universe tends toward increased entropy (randomness). Therefore, energy must be expended to bring things (e.g, molecules) into a more organized and concentrated state. Functions of active transport: 1. Provides proper chemical environment for cellular processes (e.g., ph). 2. Brings needed substrates (glucose, amino acids) &essential minerals (nitrate, phosphate, & important ions K+ and Ca++) where they are needed. 3. Gets rid of waste products (H+, Na+, C02, lactic acid). Characteristics of active transport: 1. A given systems is specific for a particular molecule or ion. 2. Transport occurs in a specific direction across membrane. Transport is accomplished by enzymes at "active sites" - i.e., substrate specific with specific orientation (directionality) in cell membrane matrix. 3. Powered by ATP molecules which "fit" into an active sites and donates high energy phosphate bonds to the process. 4. Works against continuous back diffusion (which occurs at a slower rate because it is not enzyme aided). Active transport is generally a continuous process in living cells, concentrations on either side of the membrane are maintained in "dynamic equilibrium" 2

3 2. Active Transport (cont.) 3. Mechanical: work associated with contractions Functions of contractions: 1. Locomotion 2. Organs (intestines, heart, liver, brain) 3. Cell division 4. Subcellular movements in membranes of organelles Ciliar and flagellar motion results from the coordinated sliding of outer doublet microtubules relative to their neighbors, driven by ATP. 3

4 4. Biosynthesis: creation of complex organic molecules from simple precursors, minerals, and nutrients. Synthesis is a continuous active process because molecules are constantly breaking down and need to be replaced ('Homeostasis" = maintenance). Mechanism: Biosynthetic reactions involve enzymes (proteins) in which terminal phosphates of ATP are used, directly or indirectly, to activate building-block block molecules so that they react with other building blocks in an energetically favorable manner. IMPORTANT MACROMOLECULES - precursors & functions Precursors Macromolecule Function Amino Acids Proteins Structure membranes Synthesis enzymes Active Transport enzymes Contraction - actomyosin Monosaccharides (sugars) Polysaccharides Structure membranes, cellulose (plant), chitin (animals) Carbohydrates Storage - starch (plants), glycogen (animals) Fatty Acids Lipids Structure membranes Storage - fats, oils, waxes Light absorption pigments Growth regulation - hormones Mononucleotides DNA & RNA Genetic info.- DNA Protein synthesis - RNA ATP pool (E. coli) = 1,000,000 molecules ATP utilization rate = 2,500,000 molecules/sec Turnover rate of ATP pool = 0.4 sec 4

5 Bacteria invest a large fraction of their energetic budget in proteins. Relative to some eukaryotes, E. coli is a simple organism with an extreme life-history strategy - to reproduce as fast as possible. In more complex organisms, we expect more of an energetic investment in "adaptations": a. differentiation and specialization of tissues and appendages b. metabolic or reproductive storage products Greater specialization and complexity brings the need for more information (organization). Biosynthesis is not simply the process of building molecules, but involves the organization of molecules into a hierarchy of increasing complexity: -ORGANISM -TISSUES & ORGANS -CELLS -ORGANELLES (chloroplasts, mitochondria, nuclei) -SUBCELLULAR COMPLEXES (membranes, enzyme systems) -MACROMOLECULES (proteins, carbohydrates, lipids, nucleic acids) -BUILDING BLOCKS (sugars, amino acids, fatty acids, mononucleotides) -INORGANIC PRECURSORS (CO 2, H 2 O, NH 4 ) Application of Bioenergetics to Biological Oceanography Biochemical parameters indicative of stock sizes and process rates dc 1 /dt = growth Standing Stock C 1 transfer efficiency dc 2 /dt C 2 1. MEASUREMENT OF STANDING STOCKS: Carbon = the "standard unit" of measurement, basic to all organic molecules. Bulk measurements straightforward, but difficult to separate community components: "dead" carbon (detritus) >> living C [cells] biovolume / cell carbon = population carbon microscope work is tedious carbon / biovolume varies with nutritional status, taxa, etc. 5

6 [Chl a] C:Chl a = phytoplankton carbon C:Chl varies with nutritional state, light range 25:1 (high nutrient, low light) - 200:1 more typical :1 [ATP] C:ATP = living carbon C:ATP 250:1, varies with nutritional state, taxa, etc. Other bulk measurable possible: lipopolysaccharide (LPS) bacterial cell wall component taxa-specific photosynthetic accessory pigments DNA (specific) Holm-Hansen 1969 L/O 14: Holm-Hansen and Paerl 1972 Mem. Ist. Ital. Idrobiol. 29: Flow Cytometry: another approach to quantify biomass is to count cells (and biomass) (A) Measurement Parameters 1. LFALS: Log Forward Angle Light Scatter - cell size proxy 2. L90LS: Log 90 Light Scatter - cell size proxy 3. LIBFL: Log Integrated Blue Fluorescence - DNA content (Hoechst stain) 4. LIOFL: Log Integrated Orange Fluorescence - phycoerythrin content (cyanobacteria) 5. LIRFL: Log Integrated Red Fluorescence - chlorophyll content (photoautotrophs) (B) Applications 1. Cell enumeration heterotrophic bacteria cyanobacteria prochlorophytes photosynthetic picoeukaryotes (< 3um) photosynthetic nanoeukaryotes (3-20 um) 2. Cell sorting: group-specific 1 production 3. Prochlorococcus growth rates (cell cycle) 4. Cell Abundance (cell-specific probes) 6

7 Flow Cytometry Principles of Operation: 2. Measurement of Rate Processes: Isotope (Tracer) Methods: 14 C-CO 2 Primary Production 18 O-O 2 Respiration 32 P-PO N-NH 4 + (NO 3- ) 3 H-thymidine 3 H-adenine 3 H-amino acids 14 C-amino acids Phosphorous uptake & cycling Nitrogen uptake & cycling Bacterial growth Protein synthesis 7

8 2. Measurement of Rate Processes: other approaches primary production: oxygen evolution, CO 2 consumption, ΔpH, heat production, fluorescence, change in abundance secondary production: oxygen consumption, CO 2 production, ΔpH, change in abundance PP: light bottle/dark bottle O 2 evolution grazing - dilution experiments growth rate dilution Biochemical Indices: enzymes can be induced by need (substrate availability) assumes energy economy in production of cell components ex. RUBISCO enzyme in photosynthetic dark reaction ETS activity: King and Packard 1975 L/O 20: RNA:DNA ratio protein synthesis/biomass ~ growth digestive enzymes substrate specific 8

9 3. Relationship among cell constituents: Carbon energy content - Lipid (fat) has greater than twice the energy content per carbon molecule (or per unit mass) than protein or carbohydrate. ATP energy content - ATP content is the pool of immediately available energy. The process of energy utilization can be assessed by the turnover rate of the ATP pool. Respiratory Quotient = RQ = CO 2 evolved/o 2 used a RQ b calories/g Carbohydrate Protein Lipid (Fatty Acid) a relevant to carbon budgets, given respiration measured as O 2 utilization. Conversion from stoichiometry b relveant to energy budgets, derived from chemistry. Elemental composition of organic molecules: Carbon Nitrogen Phosphorous Carbohydrates Trace Proteins Lipids Nucleic Acids Redfield Ratio (C:N:P) (molar ratio) = 106:16:1 average of all living material 4. Energetic Efficiency no process can occur with 100% conservation of energy Respiration Given: Glucose yields 686 Kcal/mole 38 moles of ATP/mole glucose respired ATP-phosphate bond liberates 10 kcal/mole 38 ATP 10 kcal/atp = 55% efficiency 686 Kcal/mole glucose Biosynthesis Given: Heterotroph starts with amino acids Glycine (AA) = 234 Kcal/mole Peptide linkage: cost = 3 ATP = 30 Kcal/mole bond energy = 5.5 Kcal/mole Needed ATP is generated with 55% efficiency 234 Kcal/mole glycine 0.55 = Kcal/mole peptide bond About one out of every 5 assimilated glycine molecules must be fully oxidized in respiration to provide enough energy to link the other 4 molecules with peptide bonds. THEORETICAL MAXIMUM EFFICIENCY: All life processes 70% 9

10 5. Energetic Implications of Adaptations Short-term - Within physiological limits, organisms adjust biochemical systems to the external environment in a manner that promotes energetic efficiency (e.g., enzymes, C:Chl). Long-term (evolutionary) - (morphological specializations, physiological tolerances). Additional capabilities and structures have energetic costs that must be offset, in the long term, by energetic advantages (e.g., homeotherm metabolism). EXAMPLE: TEMPERATURE TOLERANCE Temperature enhances all enzymecatalyzed biochemical reactions. The enzyme systems for different species are adapted to specific temperature ranges. Growth vs. temperature for 5 unicellular algae (Eppley 1972 Fish. Bull. 70) 10