Nitrogen Metabolism and Nucleic Acids

Module 6: Nitrogen Metabolism and Amino Acid Catabolism

  • Importance of Nitrogen in Biochemical Pathways

    • Nitrogen is essential for the synthesis of amino acids and nucleotides.

    • Fixed from atmospheric N2 through various processes.

  • The Nitrogen Cycle

    • Describes the transformation of nitrogen and its compounds in the environment.

    • Key Enzymes:

    • Nitrifying bacteria utilize enzymes such as nitrogenase to fix nitrogen.

    • Other enzymes include nitrates and nitrites in various states of oxidation.

    • Metabolites involved: ammonia, nitrites, and nitrates.

    • Forms of Nitrogen Used by Plants and Animals:

    • Plants: Primarily absorb nitrate ( NO3^- ) and ammonium ( NH4^+ ).

    • Animals: Obtain nitrogen through the consumption of amino acids.

  • Haber-Bosch Process

    • Industrial method for synthesizing ammonia from atmospheric nitrogen and hydrogen.

    • Critical for fertilizer production and global food supply.

  • Nitrogenase Role in Metabolism

    • Enzyme responsible for nitrogen fixation, converting atmospheric nitrogen ( N2 ) into ammonia ( NH3 ).

    • Requires ATP and is sensitive to oxygen.

  • Catabolism of Amino Acids

    • Pathway for degrading amino acids to produce energy and metabolites feeding into the TCA cycle.

    • Key Metabolite: Urea is produced for nitrogen waste processing.

  • The Urea Cycle

    • **Components to Draw:

    • Chemical names: Ornithine, citrulline, arginine, aspartate, and urea.

    • Structures of each component.

    • Enzymes involved: Carbamoyl phosphate synthetase, ornithine transcarbamylase, arginase.

    • Cofactors/coenzymes: ATP, NADH.

    • Urea Cycle Integration with Other Pathways:

    • Connects to metabolic pathways including the TCA cycle.

    • Regulation of the Urea Cycle:

    • Controlled by substrate availability and product inhibition.

  • Essential Amino Acids

    • Definition: Amino acids that cannot be synthesized by the organism and must be obtained through diet.

    • Essential Amino Acids: Include histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, and valine.

  • Intermediates for Amino Acid Synthesis

    • Six-carbon intermediates as precursors include:

    • Oxaloacetate: Precursor for aspartate and asparagine.

    • α-Ketoglutarate: Precursor for glutamate and glutamine.

    • 3-Phosphoglycerate: Precursor for serine and cysteine.

    • Pyruvate: Precursor for alanine.

  • Synthesis of Aromatic Amino Acids

    • Key Metabolites Required for Synthesis:

    • Tryptophan (Trp), Tyrosine (Tyr), Phenylalanine (Phe), and Histidine (His) require specific biosynthetic pathways often involving shikimic acid.

  • Impact of Glyphosate (Roundup) on Plants

    • Herbicide targets the shikimic acid pathway, inhibiting the production of aromatic amino acids which is lethal to plants.

    • Animals are not affected due to their ability to obtain these amino acids through diet.

Module 7: Nucleic Acids

  • DNA Structure and Function

    • DNA comprises polymerized nucleotides; serves as genetic material encoded for proteins.

    • Transcription and Translation Processes:

    • Transcription: DNA is transcribed to mRNA.

    • Translation: Ribosomes translate mRNA into proteins.

  • Definitions to Know:

    • Nucleotide: Building blocks of nucleic acids consisting of a nitrogenous base, a sugar, and a phosphate group.

    • Nucleoside: Similar to a nucleotide but lacks a phosphate group.

    • Pyrimidine: Single-ring nitrogenous bases (Cytosine, Thymine, Uracil).

    • Purine: Double-ring nitrogenous bases (Adenine, Guanine).

    • DNA Backbone: Comprised of sugar and phosphate groups.

    • Codon: A sequence of three nucleotides coding for a specific amino acid.

    • Reading Frame: The way in which ribosomes read nucleotide sequences.

    • Splicing: Process of removing introns and connecting exons in mRNA.

    • Intron/Exon: Non-coding regions and coding sequences in mRNA.

    • Capping/Tailing: Modifications of mRNA for stability and export.

    • PCR (Polymerase Chain Reaction): Technique to amplify DNA sequences.

    • Restriction Enzyme: Enzymes that cut DNA at specific sequences.

    • RNA/DNA Polymerase: Enzymes that synthesize nucleic acids by adding nucleotides.

    • Primer: Short nucleic acid sequence that determines where DNA replication or amplification will start.

  • Base Pairing

    • Watson-Crick Base Pairing: A-T and C-G nucleotide bases pair through hydrogen bonds.

    • Ability to draw the structures and indicate hydrogen bonding between A-T and C-G.

  • Types of DNA

    • A-DNA, B-DNA, Z-DNA:

    • A-DNA: DNA under dehydration conditions, more compact.

    • B-DNA: Most common form of DNA, right-handed spiral. Native DNA form.

    • Z-DNA: Left-handed spiral, associated with transcription and gene regulation.

    • Stabilization provided by hydrogen bonds and base stacking interactions.

  • DNA Structure Detail

    • Primary Structure: Sequence of nucleotides.

    • Secondary Structure: Double helix formation; major and minor grooves created by the twisting of the DNA helix.

    • Major groove: Wider, more accessible for protein binding.

    • Minor groove: Narrower, less accessible, but still interacts with proteins.

  • RNA Types and Functions

    • Three types of RNA:

    • mRNA (Messenger RNA): Carries genetic information from DNA to ribosomes.

    • tRNA (Transfer RNA): Brings amino acids to ribosomes during protein synthesis.

    • rRNA (Ribosomal RNA): Structural component of ribosomes.

  • Lac Operon

    • Functional role in the metabolism of lactose in prokaryotes, regulating gene expression in response to glucose and lactose availability.

  • Architecture of Transcription

    • RNA polymerase enzyme transcribes DNA to synthesize RNA.

    • Transcription occurs in the 5' to 3' direction, requiring other factors such as transcription factors and a promoter.

  • PCR Process

    • Components Necessary for PCR:

    • Template DNA, two primers, DNA polymerase, nucleotides, and buffer.

    • Role of Components:

    • Template DNA: Provides the target sequence to amplify.

    • Primers: Provide starting points for polymerase.

    • DNA Polymerase: Synthesizes new DNA strands.

    • Nucleotides: Building blocks for new DNA.

    • Expected Outcome: Amplification of specific DNA sequences, allowing for analysis or cloning.

  • Nucleotide Synthesis Pathways

    • Two Distinct Pathways:

    • De Novo Pathway: Synthesizes nucleotides from simple precursors.

    • Salvage Pathway: Recycles nucleotides from breakdown products.

    • Sources for Purine and Pyrimidine Synthesis:

    • Purines: Synthesized from ribose-5-phosphate, amino acids (glutamine, aspartate), and formyl group (from tetrahydrofolate).

    • Pyrimidines: Built around aspartate and carbamoyl phosphate.

  • Purine Biosynthesis First Step

    • First Step: Catalyzed by Glutamine-PRPP amidotransferase (enzyme).

    • Identify chemical structures of substrates/products involved.

    • Regulation: Feedback inhibition from end products like AMP and GMP.

    • Enzyme inhibition can be represented in Michaelis-Menten (MM) curves showing kinetic behaviors under various conditions.

  • Role of Folate in Nucleotide Biosynthesis

    • Folate: Essential for transferring one-carbon units in nucleotide synthesis, especially in purine metabolism.

  • Final Product of Inosinate Biosynthesis

    • Inosinate (IMP) is the precursor for both AMP and GMP.

    • Synthesis pathways leading to these products should be understood.

  • Pyrimidine Biosynthesis Reactions

    • First Two Reactions include:

    • Formation of carbamoyl phosphate from glutamine and bicarbonate.

    • Further reactions generating orotate.

    • Enzyme catalyzing the committed step: Carbamoyl phosphate synthetase II.

    • Regulatory mechanisms through feedback inhibition and response to nucleotide levels.

  • Understanding Enzyme Kinetics

    • Draw and interpret sample MM curves for key enzymes involved, displaying responses to inhibitors and activators relevant to nucleotide biosynthesis.

  • Control of Gene Transcription

    • Mechanisms regulating gene transcription discussed in the context of the lac operon and beyond, emphasizing the importance of regulation for cellular processes.