Exam 3- Chapter 12 Notes: Mitochondria, Chloroplasts, and Peroxisomes

Mitochondria

• Generate the majority of cellular energy.

• Breakdown lipids and carbohydrates to produce ATP.

• Synthesize some proteins using their own genome.

• Play a critical role in the generation of metabolic energy in eukaryotic cells.

• Derive energy from the breakdown of carbohydrates and fatty acids, which is converted to ATP by the process of oxidative phosphorylation.

• Mitochondrial proteins are translated on free cytosolic ribosomes and imported into the organelle by specific targeting signals.

• Mitochondria contain their own DNA, which encodes tRNAs, rRNAs, and some mitochondrial proteins.

Structure of a Mitochondrion

• Double-Membrane System:

  • Inner and outer mitochondrial membranes.

  • Separated by an intermembrane space.

• Inner Mitochondrial Membrane:

  • Folded into cristae for increased surface area.

  • High protein content (>70%) involved in oxidative phosphorylation.

  • Impermeable to most ions and small molecules to maintain the proton gradient.

• Outer Mitochondrial Membrane:

  • Permeable due to porins, allowing free diffusion of small molecules.

• Cristae:

  • Folds on the inner membrane.

  • Increase surface area for functional efficiency.

Functional Compartments / Oxidative Metabolism

• Matrix:

  • Contains mitochondrial genetic system.

  • Enzymes for central oxidative metabolism.

  • Site of glucose and fatty acid breakdown for ATP production.

• Citric Acid Cycle:

  • Takes place in the matrix.

  • Involves oxidation of acetyl CoA from pyruvate and fatty acids.

  • Yields NADH and FADH2.

• Oxidative Phosphorylation:

  • Inner mitochondrial membrane.

  • High-energy electrons transferred to molecular oxygen.

  • Energy used to create a proton gradient for ATP synthesis.

• Pyruvate and fatty acids are imported from the cytosol and converted to acetyl CoA in the mitochondrial matrix.

• Acetyl CoA is then oxidized to $CO<em>2$ via the citric acid cycle, coupled to the reduction of $NAD^+$ and FAD to NADH and $FADH</em>2$, respectively.

• The high-energy electrons from NADH and $FADH_2$ are transferred through a series of carriers in the inner membrane to molecular oxygen, coupled to the generation of a proton gradient across the membrane.

Genetic System of Mitochondria

• Mitochondria are not static organelles; they are constantly fusing and dividing.

• One role of fusion and fission is to allow exchange of genetic material. Another role is regulating susceptibility to autophagy.

• Evolutionary Background:

  • Mitochondria are thought to have evolved from bacteria that began living inside larger cells (endosymbiosis).

  • Free-living α-proteobacteria have genomes similar to mitochondria.

• Circular DNA Molecules:

  • Resemble bacterial genomes.

  • Multiple copies per organelle.

  • Vary in size among species.

• Encoded Proteins:

  • Primarily components of the oxidative phosphorylation system.

  • Also encode rRNAs and most tRNAs for translation within mitochondria.

The Human Mitochondrial Genome

• The genome contains 13 protein-coding sequences, which are designated as components of respiratory complexes I, III, IV, or V.

• In addition, the genome contains genes for 16S and 12S rRNAs and for 22 tRNAs, which are designated by the one-letter code for the corresponding amino acid.

• The region of the genome designated “D loop” contains an origin of DNA replication and transcriptional promoter sequences.

Differences Between the Universal and Mitochondrial Genetic Codes

• Codon UGA:

  • Universal code: Stop

  • Human mitochondrial code: Trp

• Codon AGA:

  • Universal code: Arg

  • Human mitochondrial code: Stop

• Codon AGG:

  • Universal code: Arg

  • Human mitochondrial code: Stop

• Codon AUA:

  • Universal code: Ile

  • Human mitochondrial code: Met

• Note: Other codons vary from the universal code in yeast and plant mitochondria.

Mitochondrial DNA Mutations

• Mitochondrial DNA can be altered by mutations. Mutations in mitochondrial genes are associated with several diseases.

• Almost all the mitochondria of fertilized eggs are contributed by the oocyte, so germ-line mutations are transmitted to the next generation by the mother.

• Leber’s hereditary optic neuropathy, which leads to blindness, is caused by mutations in mitochondrial genes that encode components of the electron transport chain.

• Elevated levels of mtDNA defects have been observed in some patients with late-onset degenerative disease.

Protein Import and Mitochondrial Assembly

• Many of the genes required for mitochondrial function and replication are in the cell nucleus.

• Most of the proteins are synthesized on free ribosomes and imported to mitochondria as complete polypeptides.

• Proteins are targeted to the matrix by amino-terminal sequences (presequences) i.e., (15-55 positively charged amino acids) that are removed by proteolytic cleavage after import.

• Presequences bind to receptors on the mitochondria that are part of a protein complex (translocase of the outer membrane, or Tom complex).

• Proteins are then transferred to another protein complex in the inner membrane (translocase of the inner membrane, or Tim complex).

• Translocation to the inner membrane requires electrochemical potential.

Targeting and Sorting Signals of Mitochondrial Precursor Proteins

• Proteins carry amino-terminal presequences.

• Presequences contain positively charged amino acids.

• Targeting to the Tom complex in the mitochondrial outer membrane.

• Targeting to Tom Complex:

  • Presequences bind to Tom complex for translocation.

• Tom Complex Entry:

  • After Tom complex passage, presequences bind to Tim23 complex in the inner membrane.

• Transfer to Tim23 Complex:

  • Import motor complex, including Hsp70 chaperone, facilitates ATP-driven translocation.

  • Some proteins with transmembrane domains exit laterally into the inner membrane.

• Matrix Translocation:

  • Mitochondrial matrix processing peptidase (MPP) removes presequences.

• Presequence Removal:

Protein Targeting to the Mitochondrial Inner Membrane

• Some proteins with multiple transmembrane domains have internal import signals instead of presequences.

• After translocation across the outer membrane, they are bound by mobile Tim9-Tim10 chaperones, which bring them to Tim22.

• The protein is transferred laterally into the inner membrane.

• Protein targeting to the mitochondrial Matrix:

  • Some inner membrane proteins are encoded by the mitochondrial genome.

  • They are synthesized on ribosomes in the mitochondrial matrix and targeted to the Oxa translocase in the inner membrane.

  • The exit Oxa laterally to insert into the inner membrane.

Sorting of Proteins to the Outer Membrane and Intermembrane Space

• Proteins destined for the outer membrane or intermembrane space also pass through the Tom complex.

• Proteins with α-helical transmembrane domains exit Tom laterally.

• β-barrel proteins pass through Tom, are bound by Tim9-Tim10 and carried to another translocon called the SAM (sorting and assembly machinery).

• SAM mediates their insertion into the outer membrane.

Mitochondrial Lipids

• Phospholipids of mitochondrial membranes are imported from the cytosol.

• In animal cells, Sphingolipids, cholesterol, phosphatidylcholine, phosphatidylinositol, phosphatidylserine are synthesized in the ER and carried to mitochondria by phospholipid transfer proteins.

• The mitochondria then synthesize phosphatidylethanolamine from phosphatidylserine.

• Mitochondria also synthesize the unusual phospholipid cardiolipin, which contains four fatty acid chains.

• Cardiolipin:

  • Is found primarily in the inner mitochondrial membrane.

  • Improves the efficiency of oxidative phosphorylation.

Transport of Metabolites Across the Mitochondrial Inner Membrane

• Small molecule transport across the mitochondrial inner membrane is mediated by transport proteins and driven by the electrochemical gradient.

• ATP/ADP exchange is powered by the voltage gradient, exporting ATP (–4) in exchange for ADP (–3).

• Phosphate (Pi) and pyruvate are imported in exchange for $OH^⁻$ ions, driven by the pH gradient.

Peptide Domains for Targeting Different Organelles

• Chloroplast: Transit peptide (TP)

• Mitochondrion: Pre-sequence

• Nucleus: Nuclear localization signal (NLS)

• Peroxisome: Peroxisomal targeting signal(s) (PTS1 and PTS2)

Chloroplasts

• The organelles responsible for photosynthesis, are similar to mitochondria in many ways:

  • Both generate metabolic energy

  • Evolved by endosymbiosis

  • Contain their own genetic systems

  • Replicate by division

• Chloroplasts are larger and more complex.

• They convert $CO_2$ to carbohydrates; synthesize amino acids, fatty acids, and lipid components of their own membranes.

• Nitrite ($NO<em>2^–$) reduction to ammonia ($NH</em>3$), essential for incorporation of N into organic compounds.

Structure of Chloroplast

• Plant chloroplasts are large organelles (5 to 10 μm long)

• Chloroplasts are bounded by a double membrane—the chloroplast envelope.

• In addition to the inner and outer membranes of the envelope, chloroplasts have a third internal membrane system, called the thylakoid membrane

• An internal membrane system, the thylakoid membrane, forms a network of flattened discs (thylakoids), which are frequently arranged in stacks called grana.

• Chloroplasts have three membranes that divide the chloroplasts into three distinct internal compartments:

  • The intermembrane space between the two membranes of the chloroplast envelope

  • The stroma, which lies inside the envelope but outside the thylakoid membrane

  • The thylakoid lumen

• Chloroplast membranes are functionally similar to those of mitochondria.

• The outer membrane contains porins and is freely permeable to small molecules.

• The inner membrane is impermeable to ions and metabolites, which must move through specific transporters.

• The stroma contains the genetic system and metabolic enzymes, including those needed to convert $CO_2$ to carbohydrates during photosynthesis.

• In thylakoid membrane - Electron transport and chemiosmotic generation of ATP takes place

• In mitochondria, electron transport generates a proton gradient across the inner membrane, which is then used to drive ATP synthesis in the matrix.

• In chloroplasts, the proton gradient is generated across the thylakoid membrane and used to drive ATP synthesis in the stroma.

The Chloroplast Genome

• Origin from photosynthetic bacteria.

• Circular DNA molecules are present in multiple copies.

• 100-200kb, contains 150 genes.

• Organelle genome encodes for ribosomal and transfer RNA and mRNA.

• One subunit of rubisco is encoded by chloroplast DNA.

• Rubisco catalyzes addition of $CO_2$ to ribulose-1,5-bisphosphate in the Calvin cycle.

• Rubisco is critical for photosynthesis, and it is thought to be the single most abundant protein on Earth.

Genes Encoded by Chloroplast DNA

Import and Sorting of Chloroplast Proteins

• Other proteins are synthesized on free ribosomes and imported into chloroplasts as completed polypeptides.

• N-terminal sequences (transit peptides) direct translocation across the two membranes of the envelope and are then removed by proteolytic cleavage.

• Proteins with N-terminal transit peptides are targeted to the Toc complex in the chloroplast outer membrane.

• Passage through the outer membrane requires ATP hydrolysis by Hsp70 and hydrolysis of GTP by Toc proteins.

• Once through the chloroplast outer membrane, the transit peptide is passed to the Tic complex in the inner membrane.

Import of Proteins into the Thylakoid Lumen and Membrane

• Sec pathway

  • Uses ATP

  • Unfolded proteins

• Tat pathway

  • Uses proton gradient

  • Folded proteins

• SRP pathway

  • Integral membrane protein

Other Plastids

• Plastids are double-membraned organelles found in plant and algal cells, involved in photosynthesis, storage, and pigment synthesis.

• Chloroplasts are a type of plant organelles called plastids.

• All have the same genome as chloroplasts but differ in structure and function.

• Chloroplasts are specialized for photosynthesis; the internal thylakoid membrane is unique.

• Other plastids are involved in other aspects of plant metabolism. They have a double-membrane envelope but no thylakoid.

Types of Plastid

• Plastids are classified based on the pigments they contain

  • Chloroplasts

    • Contain chlorophyll

    • Site of photosynthesis

  • Chromoplasts

    • Rich in carotenoids

    • Provide red, orange, yellow pigments to flowers and fruits

  • Leucoplasts (non-pigmented)

    • Amyloplasts: Store starch

    • Elaioplasts: Store lipids

    • Proteinoplasts: Store proteins

• Proplastids can develop into chloroplasts, chromoplasts, etioplasts, and amyloplasts.

• Different plastids can also convert to other types.

• Development of plastids is controlled by environmental signals and intrinsic developmental signals.

• In the photosynthetic cells of leaves, proplastids develop into chloroplasts, but only in the presence of light.

• If kept in the dark, development of proplastids is arrested at an intermediate stage (etioplasts)

Peroxisomes

• Peroxisomes: single-membrane-enclosed organelles containing enzymes involved in many metabolic reactions.

• Peroxisomes do not have their own genomes.

• Most peroxisomal proteins (peroxins) are metabolic enzymes.

• Peroxisomes can replicate by division but can also be regenerated even if entirely lost.

• Peroxins are typical eukaryotic proteins

• The oxidation of a fatty acid is accompanied by the production of hydrogen peroxide ($H<em>2O</em>2$) from oxygen.

• The hydrogen peroxide is decomposed by catalase, either by conversion to water and oxygen or by oxidation of another organic compound (represented as $AH_2$).

Function of Peroxisomes

• Breakdown of fatty acids via β-oxidation

• Detoxification of harmful substances (e.g., hydrogen peroxide using catalase)

• Metabolism of reactive oxygen species (ROS)

• Biosynthesis of:

  • Plasmalogens (important membrane lipids in the brain and heart)

  • Biosynthesis of Bile acids (in liver cells)

• Conversion of purines to uric acid

• Photorespiration in plant cells

• Peroxisomes contain enzymes required for the synthesis of plasmalogens- family of phospholipids.

• The plasmalogen is analogous to phosphatidylcholine.

• However, one of the fatty acid chains is joined to glycerol by an ether, rather than by an ester, bond.

Role of Peroxisomes in Photorespiration

• Photorespiration (in leaves):

  • Peroxisomes work with chloroplasts and mitochondria.

  • Convert glycolate (produced during photosynthesis) into glycerate.

  • Helps recover carbon and minimize waste of photosynthetic energy.

• Lipid metabolism (in seeds):

  • In oil-rich seeds, peroxisomes (called glyoxysomes) convert stored lipids into sugars.

  • Supports early seed germination by providing energy before photosynthesis begins.

• Photorespiration occurs when Rubisco fixes $O<em>2$ instead of $CO</em>2$, producing 2-phosphoglycolate, a toxic byproduct.

• Peroxisomes convert glycolate (from chloroplasts) into glycine, a safer intermediate.

• Key steps in peroxisomes:

  • Glycolate → Glyoxylate → Glycine

  • Uses enzymes like glycolate oxidase

  • Produces $H<em>2O</em>2$, which is detoxified by catalase

• Glycine is sent to mitochondria, continuing the cycle to recover carbon.

Assembly of Peroxisomes

• Origin

  • Peroxisomes can form:

    • By growth and division of pre-existing peroxisomes

    • De novo from the endoplasmic reticulum (ER)

• Peroxins (PEX proteins) recognize these signals and help import proteins.

  • Matrix proteins (e.g., catalase, oxidases) are imported post-translationally using peroxisomal targeting signals (PTS1 or PTS2).

• Mature peroxisomes grow and divide by fission to increase number

  • Peroxisomal membrane proteins are inserted into vesicles budding from the ER.