BIOL 2021 - Cell Biology - FINAL EXAM

Major structures of the eukaryotic cell.

Major differences between animal, plant, fungi, and protist cell.

Compare and contrast between the bacterial and eukaryotic cell in terms of their cell structure.

The function of proteins. Where to find it?

The function of carbohydrates. Where to find it?

The function of lipids. Where to find it?

The function of nucleic acid. Where to find it?

All cells:

A. possess a plasma membrane.

B. have mitochondria.

C. are approximately the same size.

Which statement about DNA is false?

A. It is a double helix of two polymeric strands twisted about each other.

Which of the following would not be a good reason for studying SSU rRNA to evaluate the major branches in the evolutionary history of life?

D. It mutates very frequently

Which of the following statements are characteristics of eukaryotic mitochondria and chloroplasts?

A. Their morphology is similar to Bacteria and Archaea.

B. Both contain their own DNA. Both multiply by binary fission.

C. They contain their own transcription and translation machinery.

D. Contain an ETC on the inner membrane.

Human cells do not contain any circular DNA molecules.

True

During protein synthesis amino acids are added to a growing peptide chain. The addition of each amino acid involves which reaction?

B. Dehydration reaction

Primary Structure

→ The sequence of amino acids in a polypeptide chain.

Secondary Structure

→ Local folding patterns like alpha-helices (α-helices) and beta-sheets (β-sheets) formed by hydrogen bonding.

Tertiary Structure

→ The overall 3D shape of a single polypeptide, including interactions like hydrogen bonds, disulfide bridges, and hydrophobic interactions.

Quaternary Structure

→ The arrangement of multiple polypeptide chains (subunits) in a protein complex.

How large is a typical eukaryotic cell?

B. 10-20 µm across

What type of microscopy was used in the micrograph below?

A. Transmission EM

One tenth (0.1) of a mm =

B. 10 µm

Types of light microscopy

• Bright field

• Phase-contrast

• Differential interference

• Dark-field

• Fluorescence

Bright Field

Generates a dark image of an object over a light background

Phase Contrast

converts small differences in phase into contrast to generate an image

• exploits differences in refractive index between the cytoplasm and the surrounding medium or between different organelles.

• can be used to view live cells and cellular organelles

Dark field microscopy

• enable microbes to be visualized as halos of bright light against darkness.

• Allows the detection of objects that are unresolved by bright-field microscopy

Fluorescence Microscopy

The specimen absorbs light of a defined wavelength and then emits light of lower energy, thus longer wavelength; that is, the specimen fluoresces.

Magnification

How much larger an object appears than it is.

An increase in the apparent size of an image to resolve smaller separations between objects.

Detection

the ability to determine the presence of an object.

Resolution

Ability to see separate objects as separate

Contrast

Differences in light intensity (amplitude) between an object and its background.

How can resolution be increased in microscopy?

Different ways the contrast can be generated in microscopy.

Advantages of microscopy

Disadvantages of microscopy

Processing of tissue samples/staining

Advantages and disadvantages of fluorescence microscopy.

Electron microscope (EM)

Light microscopy

Transmission microscopy.

Scanning electron microscopy.

Is the inside of the cell ordered or disordered

Ordered

Name the organisms which is proposed to be the precursor for mitochondria?

a-proteobacteria

Name the organism which is proposed to be precursor for chloroplasts. Provide evidence.

Cyanobacteria.

Evidence:

• morphology

• Reproduction- divide by binary fission

• Genetic information

• Ribosomes, transcription and translation

• Electron transport

List a molecular marker for assembling the tree of life.

SSU rRNA

The DNA code is faithfully replicated by…

polymerases

Genes are … into discrete molecules: RNA

transcripted (transcription)

RNA is then … into proteins, which are the workhorses

transcribed (translation)

The … … delineates the cell and controls the exchange and communication with the nonliving world that surrounds the cell.

plasma membrane

TRANSCRIPTION

is the conversion of the DNA code (gene) into RNA

• RNA are intermediates of information transfer.

• Transcription permits a choice: express or repress a gene

TRANSLATION

ribosomes read mRNA into protein

Catabolism

reactions that are highly exergonic (increase disorder and/or liberate energy) – spontaneous.

Anabolism

reactions that are endergonic (decrease disorder and/or “store” energy in bonds) – not spontaneous

ATP (adenosine triphosphate)

A common energy “currency” molecule. Carried by the activated carrier molecule

Prokaryotes

• lacks a nucleus enclosing the genetic information (DNA)

• lacks other membrane-bound organelles.

• The plasma membrane is the only membrane structure of these cells.

• are mainly bacteria and archaea

Eukaryotes

• Genetic information (DNA) is enclosed by the nucleus, a membrane-bound organelle

• Contain various membrane-bound organelles with distinct functions

• Contain a cytoskeleton

• Mitochondria and chloroplasts are organelles with their own genome

Domain of life

Bacteria, archaea, eukaryotes

LUCA

Last universal common ancestor

LUCA of all living organisms

• Lived about 3.5 million years ago

• Was a complex cell

• Had enzymes that could synthesize lipids to make cell membranes

• Had complex metabolic enzymes and pathways

• Had translation (and ribosomes)

phagocytosis

The process by which cells eat other cells (e.g. white blood cell, neutrophil, eating a red blood cell)

Plausible Origin of Eukaryotic Cells (NUCLEUS)

• Primordial eukaryotic cells may have been predatorial, eating other cells

• White blood cell, neutrophil, eating a red blood cell by phagocytosis

• Requires change in cell shape driven by the cytoskeleton filaments

• Likely, nuclear enclosure would be advantageous to protect DNA from entanglement and breakage

• Predatorial eukaryotic cells may also explain the origin of mitochondria and chloroplasts

Mitochondria and chloroplasts (similarities)

• Energy organelles

• Are defined by a double lipid layer (like the nucleus)

• Have their own genome.

• Have their own ribosomes and transfer RNAs to make proteins (like cells)

The name of the bacterium involve in the Plausible Origin of Mitochondria

aerobic prokaryotic alpha-proteobacteria (alpha-proteobacteria)

Plausible Origin of Mitochondria

• Predation of oxidizing bacteria

• Became symbiotic: large cell provided protection and food molecules and bacteria oxidized food to release energy

• Symbiotic relationship eventually became permanent by loss of redundant genomes

The name of the bacterium involve in the Plausible Origin of Chloroplasts

cyanobacteria

Plausible Origin of Chloroplasts

• Predation of photosynthetic bacteria (cyanobacteria)

  • Prochlorococcus-Synechococcus type

• Became symbiotic: cyanobacteria converted sunlight to food while large eukaryotic cell with mitochondria oxidized food molecules to chemical energy

• Symbiotic relationship eventually became permanent by loss of redundant genomes

ATP (adenosine triphosphate)

• A common energy “currency” molecule

• Carried by the activated carrier molecule

Plausible Origin of Eukaryotic Cells (Stages)

• Stage 1:

  • Ancestral pre-eukaryotic cells were likely predatorial, eating other cells by phagocytosis.

    • Evolved the nucleus to protect DNA molecules from damage

• Stage 2:

  • Predation of oxidizing bacteria leads to endosymbiosis + eventually mitochondria

• Stage 3:

  • A line of mitochondria-containing eukaryotic cells may have internalized photosynthetic bacteria, which became chloroplasts.

Proteins

• Made of amino acids linked by peptide bonds.

• Function as enzymes, structural components, transporters, and signaling molecules.

• Example: Hemoglobin (oxygen transport), enzymes (catalyze reactions), actin (cytoskeleton support).

Nucleic Acids

• Include DNA (stores genetic information) and RNA (involved in protein synthesis).

• Made of nucleotides (A, T/U, C, G).

• Example: mRNA (carries genetic instructions), tRNA (helps in translation), ribosomal RNA (forms ribosomes).

Lipids (Fats & Membranes)

• Hydrophobic molecules that make up cell membranes and store energy.

• Include phospholipids, cholesterol, and triglycerides.

• Example: Phospholipids (form the lipid bilayer), cholesterol (membrane fluidity), triglycerides (energy storage).

Carbohydrates (Sugars & Energy)

• Made of monosaccharides (simple sugars like glucose).

• Provide energy and structural support.

• Example: Glucose (energy source), glycogen (stored energy in animals), cellulose (plant cell walls).

Macromolecules

The shape, size and physical and chemical properties of macromolecules allow them to have specific functions.

Monomers

= twenty different amino acids

Macromolecules Covalent linkage and rearrangement change by…

• 1. Polymer Length 

• 2. Linear sequence of Monomer Types

• 3. Properties of the Covalent bonds between monomers , i.e., where on a molecule does one attach the next building block – important for polysaccharides

• 4. Conformation DNA of a molecule, i.e., 3-D shape of a molecule

ligands

Interacting molecules can be small molecules or other macromolecules

bind to proteins via noncovalent bonds

bind to specificity and strength (affinity) depends on type and number of non-covalent bonds

Limit of Resolution

The smallest distance by which two points can be separated yet still be resolved as two points.

NA = n sin θ

λ = wavelength of light (μm)

NA = Numerical Aperture (measure of light gathering power of lens)

As d gets smaller, resolving power increases

Processing of tissue samples is required for staining

Fixation, Embedding, and Sectioning

• Fixation: covalent cross-linking locks proteins into place.

• Embedding: wax permeates and then solidifies to harden and stabilize tissue.

• Sectioning: tissue is thinly sliced with a microtome for observation under a light microscope.

Fixation

covalent cross-linking locks proteins into place.

Embedding

wax permeates and then solidifies to harden and stabilize tissue.

Sectioning

tissue is thinly sliced with a microtome for observation under a light microscope.

Phase Shifts in Light

in phase and out of phase

Bright Field

Generates a dark image of an object over a light background

Phase Contrast

Visualize living cells not all structures are visible

Phase Contrast (differences)

exploits differences in refractive index between the cytoplasm and the surrounding medium or between different organelles.

can be used to view live cells and cellular organelles

Dark field microscopy

Dark-field optics enable microbes to be visualized as halos of bright light against darkness.

Allows the detection of objects that are unresolved by bright-field microscopy

Which type of light microscope do you think would be most useful and why?

Limitations of light microscopy

Solutions: dyes, fluorescent probes, digitization, different types of microscope

fluorescence

Electrons can absorb photons and move to an excited state.

Excited electrons can release energy as light when they return to the ground state:

Fluorescence Microscopy

The specimen absorbs light of a defined wavelength and then emits light of lower energy, thus longer wavelength; that is, the specimen fluoresces.

natural fluorescent (auto fluorescence)

e.g. chlorophyll

How do you make cells/molecules fluorescent?

Light microscope, dark field, bright field, phase contrast, DIC, fluorescence

Resolution magic number 200 nm

Fluorophore

fluorescent chemical compound

Its cell specificity can be determined by:

chemical affinity

labeled antibodies

dna hybridization

fusion reporter

Immunocytochemistry

primary antibody: rabbit antibody directed antigen A

secondary antibodies: marker-coupled antibodies directed against rabbit antibodies

antibody binds specifically to the antigen

Immunocytochemistry (downside)

usually cells must be fixed aka dead, permeabilized and often partially extracted

fixed cells provide only a snapshot of what is going on or present in the cell

Green fluorescent protein (GFP)

Chromophore responsible for GFP fluorescence is shielded from quenching by aqueous solvent

ex. Serine-tryrosine-glycine

The gene for GFP can be inserted and expressed in prokaryotic and eukaryotic cells

The gene for GFP can be fused with other genes to study the expression and localization of specific proteins

Green fluorescent protein (GFP) is a bioluminescent protein isolated from jellyfish aequorea victoria

transmission electron microscope (tem)

transmission EM allows you to visualize molecular structure

high resolution (~1nm)

see lots of internal structure and molecular structure

BUT – requires thin sections (25-100nm) ~200 to go through a cell

TEM (downsides)

cell fixed

embedded in resin

Sectioned

stained with electron dense material

just see where the stain sticks

Immunogold Electron Microscopy

Detect specific molecules (like immunocytochemistry)

Stain thin slices with primary antibody against protein of interest

Follow with secondary antibody conjugated with gold-particles

Gold-particles are electron dense and can be 5-20 nm in size.

Can do dual staining using different secondary antibodies conjugated with different sized-particles

Scanning Electron Microscopy (SEM)

surface is coated with heavy metal

measures quantity of electrons scattered over surface