BIOL Lecture Outline Chapter Metabolism The Use of Energy

BIOL 3702 Lecture Outline Chapter 10: Metabolism: The Use of Energy in Biosynthesis Principles of Biosynthesis ◆ Anabolism (= biosynthesis) ✳ Creation of more complex molecules from simpler ones, but requires energy input ✳ Carefully balanced with catabolic processes ✳ Energy required in terms of ATP is enormous ◆ Biosynthesis is shaped by several guiding principles: ✳ All macromolecules are derived from about 30 small precursors ✳ Many of the same enzymes used in both catabolism and anabolism ✳ End product regulation is generally more important in anabolic pathways ✳ To be efficient, anabolic pathways must operated irreversibly and are usually coupled with the breakdown of ATP ✳ In eucaryotes, compartmentation permits different pathways to operate simultaneously and independently ✳ Anabolic and catabolic pathways typically use different co-factors Precursor Metabolites ◆ Anabolism requires precursor metabolites (carbon skeletons) to synthesize monomers ◆ Precursor metabolites are intermediates of the glycolytic pathways and TCA cycle ◆ These pathways are often referred to as central metabolic pathways and are important to both heterotrophs and autotrophs Photosynthetic CO2 Fixation ◆ Most microbes can incorporate, or fix, CO2 via anaplerotic or other pathways ◆ Only autotrophs use CO2 as a sole carbon source and, therefore, utilize one of three different reductive pathways to do so ◆ Energy for these pathways comes from photosynthesis or lithotrophy Page 1 of 5 BIOL 3702 Lecture Outline Chapter 10 ◆ Three pathways to fix CO2 ✳ Calvin Cycle (reductive pentose phosphate cycle) ● ● Present in photosynthetic eucaryotes and most photosynthetic procaryotes Absent in Archaea, some procaryotic obligate anaerobes and microaerophiles ✳ Reductive Tricarboxylic Acid Pathway - used by some Archaea and anoxygenic phototrophs ✳ Acetyl-CoA Pathway - used by acetogens, methanogens, and sulfate reducers ◆ Calvin Cycle ✳ Occurs in: ● ● Chloroplast stroma (eucaryotes) Carboxysomes (procaryotes) Carboxylation – ✳ Three phases ● CO2 is incorporated into ribulose 1,5-bisphosphate (RuBP; 5 carbon molecule) to form two molecules of 3-phosphoglycerate (PGA; 3 carbon molecules) Catalyzed by the enzyme ribulose-1,5-bisphosphate carboxylase PGA is reduced to glyceraldehyde 3-phosphate Requires one molecule of ATP and one molecule of NADPH for this reduction process to occur RuBP is regenerated via a series of reactions that resemble the pentose phosphate pathway Also generates a number of different carbohydrate structures that can be used elsewhere in the cell – ● Reduction Phase – – ● Regeneration Phase – – ✳ Total energy expenditure (provided by photosynthesis or chemotrophy): ● ● 2 ATP molecules 3 NADPH molecules Gluconeogenesis ◆ Non-photosynthetic microbes synthesize sugars from reduced organic molecules ◆ Synthesis of glucose using noncarbohydrate precursors is termed gluconeogenesis ◆ Gluconeogenic pathway is remarkably similar to reverse glycolysis Page 2 of 5 BIOL 3702 Lecture Outline Chapter 10 ◆ Gluconeogenic pathway ✳ Shares seven (7) enzymes with glycolysis ✳ Three steps are irreversible ◆ Some sugars are synthesized as nucleotide diphosphates which play a role in polysaccharide formation Amino Acid Synthesis ◆ Amino acids arise ✳ Assimilation reactions involving nitrogen and sulfur ✳ From carbon skeletons that are intermediates in many common pathways, e.g., methionine is derived from oxaloacetate formed in the TCA cycle ◆ To make amino acids, microbes require large amounts of phosphorous (P), sulfur (S), and nitrogen (N) ✳ P, S, and N comprise various portions of organic molecules used by cells ✳ Each of these elements is assimilated (incorporated) into organic molecules using different pathways ◆ Phosphorous assimilation ✳ Common sources ● ● Inorganic phosphate Organic phosphate esters Photophosphorylation Oxidative phosphorylation Substrate-level phosphorylation ✳ Inorganic phosphate is obtained through the formation of ATP by: ● ● ● ✳ Organic phosphate is obtained by microbes from hydrolyzing organic phosphates esters with enzymes termed phosphatases ◆ Sulfur assimilation ✳ Microbes obtain sulfur from two sources: ● ● Sulfur-containing amino acids (cysteine and methionine) Sulfate (SO4-) by assimilation sulfate reduction pathway - SO4- is converted to H2S which is then incorporated into cysteine Page 3 of 5 BIOL 3702 Lecture Outline Chapter 10 ◆ Nitrogen is acquired by assimilation or fixation ✳ Assimilation reactions ● Ammonia (NH4-) assimilation – – Direct incorporation of NH4- into organic material to form amino acids Indirect formation of amino acids by transamination reactions ● Assimilatory nitrate (NO3-) reduction - NO3- is reduced to nitrite (NO2-), which is then further reduced to NH4- and subsequently incorporated into amino acids ✳ Nitrogen fixation - atmospheric nitrogen (N2) is catalyzed by the enzyme nitrogenase in a three step reduction process to form NH3, which can be subsequently used to form amino acids Some amino acids are made via transamination reactions Such processes conserve cellular energy and materials Synthesis of amino acids tends to be highly regulated ◆ Amino acids biosynthetic pathways ✳ ✳ ✳ Anaplerotic Reactions ◆ The demand for intermediates of pathways can lead to a depletion of these compounds, thus disrupting the particular metabolic process, e.g., use of oxaloacetate from the TCA cycle to make amino acids ◆ Microbes have developed reactions, termed anaplerotic reactions, to replace these lost intermediates ◆ Most microbes will replace TCA cycle intermediates by CO2 fixation - NOT the same as the Calvin Cycle or other pathways that fix CO2 to produce carbohydrates ✳ CO2 is added to another compound that forms an intermediate ✳ Functions as a replacement process, not for cellular growth ✳ Maintains the operation of the TCA cycle ◆ Some microbes use the glyoxylate pathway to maintain the operation of the TCA cycle as well as to grow on two-carbon sources (e.g., acetate) ✳ Essentially a modified TCA cycle ✳ Two key enzymes: ● ● Isocitrate lyase Malate synthase ✳ Pathway is essential for the survival of some pathogenic microbes in vivo Page 4 of 5 BIOL 3702 Lecture Outline Chapter 10 Nucleotide Synthesis ◆ Purines and pyrimidines are cyclic nitrogenous bases that are crucial to cellular function ✳ Pyrimidines - cytosine, thymine, uracil ✳ Purines - adenine and guanine ◆ Pyrimidines and purines form nucleosides when combined with a ribose or deoxyribose sugar ◆ Nucleotides are nucleosides with one or more phosphate groups attached, e.g., ATP, ADP, etc. ◆ Purine biosynthesis ✳ Inosinic acid produced from ribose 5-phosphate (11-step sequence) involving the cofactor folic acid [folic acid production is inhibited by sulfonamide drugs] ✳ Inosinic acid is metabolized to adenosine and guanosine monophosphates Lipid Synthesis ◆ Fatty acid synthesis uses a fatty acid synthase and an acyl carrier protein to complex together acetyl-CoA and malonyl-CoA into long chain molecules ◆ Triacylglycerols are produced from fatty acids and glycerol phosphate using a phosphatidic acid intermediate ◆ Phospholipids are made from phosphatidic acid via a CDP-diacylglycerol intermediate combined with an amino acid Page 5 of 5