BSC360: Exam 2 Information

In vitro

In vitro

•  experiments performed with cellular structures or tissues in a test tube 

  • Outside the organism 

  • Ex: cell culture: the process of growing cells isolated from an organism in a cell culture dish 

    • Need:

      • Medium that contains essential nutrients and is at physiological pH 

      • Humidity 

      • Physiological temperature 

      • Growth factors 

Adherent cells

• Cells that have to attach to a solid surface; most animal cells 

Suspension cells

• cells that can float around solution; bacteria, yeast, and some animal cells 

Primary Cells

• cells isolated from tissues 

  • Can be cultured in dishes for several generations 

  • Limited life span 

  • Grows in a monolayer 

  • Contact inhibition: stop dividing when cell contact each other

Cell lines

• derived from GMO cells; isolated from tumor tissues 

  • Transformed cells; immortalized 

  • Grow indefinitely 

  • Rapid growth rate

  • Lost contact inhibition 

  • Tumor morphology 

In vivo

• experiments conducted in living organisms

  • Ex: clinical trial, drug testing 

Reverse Transcription

• uses the enzyme reverse transcriptase to make DNA from an RNA template 

  • mRNA is pureed from cells of interest 

  • poly -T primer binds to poly-A tail of mRNA; makes it DS 

    • Reverse transcriptase needs DS nucleotide to bind and begin transcription 

  • Reverse transcriptase uses mRNA template to make cRNA 

  • RNA is partially degraded by RNases 

    • DNA polymerase needs DS nucleotide to bind and begin transcription 

  • DNA polymerase uses cDNA strand as a template to make DS cDNA 

  • Result: complementary DNA strand containing DNA copy of mRNA 

Polymerase Chain Reaction

• Amplification of small parts of DNA in a test tube 

• Uses DNA polymerases found in thermophilic bacteria bc of its ability to survive high heat needed to denature protein 

• Exponential reaction: initial copy becomes template for other DNA segments 

• DNA is heated to melting temperature-> breaks hydrogen bonds between strands and allows them to separate 

  • Becomes SS

• Sample is cooled just enough to allow primers to bind to their specific complementary section of DNA 

• DNA synthesis: 

  • Sample is heated to the working temperature of DNA polymerase 

  • Polymerase binds to the DNA primer complex and synthesis of a new DNA strand 

• PCR rxn continues-> by third cycle you have DNA that is only from your area of interest 

  • DNA segment becomes predom further in the cycle 

  • The cycle runs 30x times

Fluorophore

molecule with fluorescent chemical that emits wavelengths when light hits it

absorbs light energy→ light absorpotion excitation of electrons ->the fluorophore re-emits the absorbed light energy at a larger wavelength upon the electrons return to their basic state 

DAPI

DNA dye that is exicted by 354 nM (UV LIght) and emits a blue light

Phalloidin

dye Toxin that binds to actin filaments; Can be conjugated to fluorescent dye 

Phosphatidylserine

• Most abundant negatively charged phospholipid 

• In healthy cells: found only on the cytosol side 

  • If on the extracellular side-> signal for apoptosis

Phosphatidylcholine

• Most abundant phospholipid in eukarytoes 

• Most commonly found on the extracellular side on the membrane

Sphingomyelin

• Regulator of cholesterol distribution within membranes 

  • Involved in cholesterol homeostasis

Phosphatidylethanolamine

• Helps fold certain membrane proteins 

• Essential for mitochondrial respiratory complexes 

• Plays a role in autophagy (cell cycling) 

Micele

type of bilayer formation. Cosists of a core and a shell where hydrophobic end groups form the core and hydrophilic head groups form the outer shell

Cholesterol

• Contains rings of hydrocarbon structures in the hydrophobic part 

• Hydrophilic part: hydroxyl group 

• Abundant in mammalian cells 

• Absent in prokaryotes 

• Affects the membrane permeability and mobility of other membrane lipids 

• Membrane lipid-> precursor of steroid hormones 

Glycolipids

• Sugar group in hydrophilic heads 

• Hydrophobic fatty acid tail 

• Found on the surface of all plasma membrane (non cytosolic side)

cold

temperature in bilayer that results in phospholipids pack in tighlty making membrane rigid and inflexible; gel like consistency

→ Cholestrol molecuels counteract this

hot

the temperature of  phospholipids where are loosely packed making membrane flexible, un able to hold shape; fluid-like consistency 

Cholesterol: holds phospholipids together stopping them from becoming loose: increases rigidity

unsatured fatty acid tails

•  kinks in their hydrocarbon tail 

  • Harder to pack; increases fluidity 

  • Harder to freeze

  • Creates small spaces for small molecules allowing certain small molecules to easily cross the membrane

organelles without membranes

• Ribosome 

• Centrosome (structure that organizes mitotic fibers) 

• Cytoskeleton: microfilaments, microtubules

Phospholipids

• Phosphate group with hydrophilic head 

• Hydrophobic Fatty Acid Tails: 

  • Can be unsaturated: results in a kink in the tail 

• Asymmetrically distributed into two leaflets of the bilayer

• Most abundant membrane lipids 

Fluid Mosiac Model

dynamic and flexible structure of membranes made up of proteins, phsoplipids, glycolipids and cholestrol

Transmembrane proteins

types of integral protein

• Span the lipid bilayer by alpha helical segments

• ThreForms: alpha-helical proteins, and beta barrels with hydrophobic side chains

Monolayer associated

protein located almost entirely in the cytosol

→ amphipathic helix is snug in the bilayer

Lipid anchored proteins

• Linked to one or more lipid molecules through different covalent bonds 

• Protein itself does not enter the bilayer

Peripheral Proteins

• proteins that do not interact directly with the lipid bilayer

  • Associate with the membrane by interaction w/ transmembrane proteins or hydrophilic heads of lipids 

  • Locaized at either side of the bilayer 

mRNA

type of rNa that carries the nucleotide sequence that is used as the recipe for protein translation

rRNA

type of RNA that is resposbnible for ribosome cataylticc activty and function

tRNA

type of RNA that carries amino acids to the ribosome for addition into the growing polypeptide chain

siRNA and miRNA

RNA that greulates protein translation by causing mRNA degradation

Wobble Hypothesis

a theory that states that the first two nucledtides of a codon are more important for tRNA binding during translation

ribosome

organelle not encoloded in a membrane

channel proteins

proteins that transport small hydrophilic molecules ions or H20, involved in passive transport, form a hydrophilic passage, and transport molecules down the concentration gradient

ex: K+ Channel

hypotonic, hypertonic

osomis is the transportaion of water across a semi[ermable membrane from a __________ solution to a ______ solution

Cholesterol

membrane lipid that does not contain a fatty acid tail

Passive Transport

With the concentration gradient - moving a solute from an area of high concentration to an area of low concentration

Active transport

Against the concentration gradient - moving a solute from an area of low concentration to an area of high concentration

thylakoid membrane

where are the proteins of the elctron transport chain located in chloroplasts?

d. creates a H+ graident that ATP synthase uses to create ATP

what is true for all electron transport chains

a. cytochrome b6-f pumps H⁺ across the membrane that ATP synthase will utilize to create ATP

b. Water is a product of all electron transport chains

c. NADH is the high energy electron carrier

d. Creates a H⁺ gradient that ATP synthase uses to create ATP

stroma

light indepedent reactions produce sugar and occurs in the

ATP, NADPH, oxygen

light dependent reactions products

CO2, H20, and ATP

the oxidation of food molecules produces

c. ATP synthase inhbitor

Electron transport is coupled to ATP synthesis in mitochondria and in chloroplasts. Which of the following is likely to affect the coupling of electron transport to ATP synthesis in all these systems?

a. the absence of light

b. a potent inhibitor of cytochrome c oxidase

c. ATP synthase inhibitor

d. the removal of oxygen

pyruvtae, ATP, NADH

final products of glycolysis

Transporter (carrier proteins)

• Interacts with a molecule to be transported, like an enzyme interacting with its substrate

• Confromational changes

• UNIPORTER:

• SYMPORTERS:

• ANTIPORTERS:

partipates in passive transport

uses free energy released by the movement of one molecule down its cocnentration graident to power the movemnet of the other molecule aganist its concentration graident

UNIPORTER

• type of transporter protein, single type of molecule moved down its concetrationg gradient

  • Bind site shifts from outside to inside positon  

SYMPORTERS

type of transporter protein,  two types of molecules in the same direction; coupled transporters 

antiporters

type of transport protein,  two types of molecules to opposite directions; coupled transporters

coupled transport

occurs in transport proteins (usally antiporters and symporters ) Uses free energy released by the movement of one molecule down its concentration gradient to power the movement of the other molecule against its concentration gradient 

Pump transporter

type of transproter that partipates in active transport

ex: ATP powedered pumps (ATPases) that uses the enerfy from hydrolysis of ATP to pump out Na+

electrochemical gradient

•  the concentration gradient of the solute and a force from the memrabne potneital 

  • Voltage and concentration work in opposite directions: Membrane potential makes transport of solute harder

polar and ions

what type of molecules require transport proteins to help them move across

cristae

inner membrane folds of the mitochondria that contains and organizes the elctront ransport chain and ATP synthase

→ carries out oxidative phsophoroylation

→ allows H+ graident formation by seperating matrix from inner membrane spave

3 NADPH, 1 GTP, 1 FADH2, 2 CO2 Released

net result of the citric acid cycle

Chemiosmosis

• The movement of ions across a semipermeable memraben bound structure down their electrochemical gradient 

• occurs during oxidative phosphorylation H+ gradient drives ATP synthase to produce ATP:

  • ADP +Pi -> ATP,

32- 34 ATP, H2O

final products of the oxdiative phosphorlation

38

max amount of ATP that can be produced through cellular respiration with one glucose molecule

Acetyl CoA

pyruvate is converted into this molecule in the mitochondrial matrix

releases CO2 and converts NAD+ to NADH

cytosol

glycolysis occurs in what part of the cell

mitochondrial matrix

the citric acid cycle occurs in what part of the cell