BSCI330 Final Exam

P-Type Pump

use of a high energy phosphoprotein intermediate, typically an ion pump where ATP is hydrolyzed and attached to the pump, the phosphate attachment leads to conformational change

F-Type Pump

found in membranes of mitochondria, bacteria, chloroplasts; generates ATP through use of H+ gradient

V-Type Pump

found in membranes of lysosomes, synaptic vesicles, plant vacuoles; regulate the pH environment by pumping H+ into these compartments

ABC Transporter

ATP Binding Cassettes that typically pump small molecules rather than ions, multiple domains, typically pump hydrophobic molecules up a gradient and across a membrane

Cytoskeleton Major Components (Size Order)

microtubules > intermediate filaments > microfilaments

Microtubules

polymers of tubulin

Intermediate Filaments

polymers of helical proteins

Microfilaments

polymers of actin

Polymerization of Cytoskeltal Monomers

requires mucleoside triphosphates in the form of GTP (tubulin) or ATP (actin); cytoskeletal monomers containing NTP have higher affinity for binding partners than those w/ NDP; polymerization at + end, depolymerization at - end; NTP bound monomer at + end, NDP bound monomer at - end

Actin

composed of a network of flexible filaments dispersed throughout a cell - highly concentrated beneath plasma membrane; form basis of cell shape and structure; form contractile rings of dividing cells; aid in contraction of muscle cells; propel vesicles and other cellular components through cytoplasm; soluble, globular protein

Polymerization of Actin

actin monomers bound to ATP are added to + end of growing filament, actin ADP monomers are lost from depolarizing - end

Dynamic Instability “Treadmilling”

addition at + end is equal to removal at - end

Rho Family of GTPases

act as molecular switches to control actin polymerization dynamics by regulating the activity of actin-binding accessory proteins; Rho-GTP, Rac-GTP, Cdc42-GTP

Rho-GTP

actin bundling, generates stress fibers (bundled fibers containing actin/myosin)

Rac-GTP

actin polymerization, generates lamellipodia (sheet-like plasma membrane projections) and membrane ruffles

Cdc42-GTP

actin polymerization and bundling, generates filopodia (tube-like plasma membrane projctions) and short cell protrusions called microspikes

Microtubules

form a network of rigid tubules that radiate through the cytoplasm of all eukaryote cells, mitotic spindles of dividing cells, the core of motile appendages: cilia and flagella; formed from alpha and beta tubulin; diverse family of soluble, globular proteins

Polymerization of Tubulin

\+ end of microtubules grows by addition of tubulin dimers bound to GTP, GTP hydrolyzes to GDP, - end of microtubules contains more GDP and grows more slowly (tubulin dimers lost from this end)

GTP Cap

tubulin GTP-dimer at the + end that forms since the rate of polymerization at the + end is more rapid than the rate of GTP hydrolysis

Catastrophe

GTP cap lost and plus end undergoes rapid depolymerization if the rate of GTP hydrolysis exceeds the rate of polymerization

Microtubule Regulation - Stability

growth or shrinkage of microtubules can be regulated by altering the balance between addition and removal of tubulin dimers

Microtubule Regulation - Orientation

assembly happens in 2 phases; nucleation (small portion of tubule formed at beginning) and elongation (addition of tubulins and GTP-cap); microtubule organizing centers (MTOC’s) play a role in nucleation

Centrosomes

MTOC’s found in animal cells that divide right before cell division begins

Proteins as Molecular Motors

shape changes in proteins generates movement, coupling shape change in one direction with ATP hydrolysis makes movement favorable

Myosins

huge family of motor proteins that bind to actin microfilaments

Myosin II

heteromer with 6 polypeptide chains - one pair heavy and two pairs light; molecules can associate into filaments that are highly stable in muscle cells and form basic structural unit of contractile machinery; only formed transiently in non-muscle cells

Kinesins

microtubule-associated motor proteins that move vesicles and organelles along nerve axons from cell body to the synaptic terminals; movement only occurs from the - end to the + end

Adaptor Proteins

binds cargo to the kinesins

Dyneins

microtubule-associated motor proteins that can bind and transport cargo via its light chains; movement only from the + end to the - end

Intermediate Filaments

composed of a group of related long helical proteins, provide mechanical strength, not in every cell type, form via coiled-coil interactions of alpha-helical proteins, first form dimers which assemble in staggered fashion to form ropelike filaments

Semi-Conservative Replication

each DNA strand is used as a template for the synthesis of a complementary strand

Replication Direction

5’ → 3’, new nucleotides are added at 3’ end, chain growth occurs 5’ to 3’, addition of deoxynucleotides requires a primer

DNA Polymerase

enzyme with fingers, palm, and thumb domains that catalyzes DNA replication

Solution for Strand Polarity

all synthesis 5’→3’, Okazaki fragments are synthesized on lagging strand, DNA ligase seals gap between successive fragments

Solution for Unzipping DNA

DNA helicase unzips DNA and uses ATP to act as a rotary engine, unzipped DNA is stabilized by single-stranded DNA binding protein

Solution for Processivity

sliding clamp holds DNA polymerase in place, clamp loaded on DNA by clamp loader that uses ATP hydrolysis to lock clamp around DNA

Topoisomerase

relieves torsional stress in DNA, prevents supercoiling, solves issue of DNA unwinding

Topoisomerase 1: Nick and Swivel Mechanism

introduces a nick into one strand of the DNA, two strands of DNA can now rotate relative to each other at the point of the nick to relieve accumulated strain, restores bond of cut strand without the use of ATP

Topoisomerase 2: Gating Mechanism

binds to double-stranded DNA and makes a double-stranded cut, one portion of DNA passes through the gate of the enzyme, reseals the DNA using ATP

Replication Origin

region of high AT content where DNA replication begins

Solving for One Copy (Prokaryotes)

A in origin GATC sequence is methylated, origin has a refractory period when only one strand is methylated, replication can only occur when both strands are methylated, mediated by Dam methylase

Solving for One Copy (Eukaryotes)

pre-replicative complex includes a helicase and assembles on origin in G1 phase, pre-RC is held inactive by proteins until phosphorylation by a cell division kinase (cdk) during S phase, activation by cdk durin S phase results in phosphorylation of origin replication complex (ORC) and activation of helicase, phosphorylated ORC keeps oriin inactive once replication has completed until after M phase

Histone Replication

H3-H4 tetromers are randomly assorted between the two strands that are being replicated, H2A-H2B dimers and new H3-H4 tetramers are loaded using histone chaperones, re-establishment of histone marks after cell division is crucial for the maintenance of gene expression programs

“End-Replication” Problem

can lead to progrssive DNA loss from the ends of linear chromosomes, 5’ end of eukaryotic linear chromosomes cannot be replicated by standard mechanism

Telomerase

prevents linear DNA ends from being lost during replication by forming telomeres

Solving for Accuracy Problem

DNA polymerase can proofread due to dependency of perfect base pairing of the -1 base

Strand-Directed Mismatch Repair

newly synthesized strand recognized by presence of an unsealed nick, re-synthesis after elimination of a section of the mismatched strand fixes the error

Catabolic Reactions

extract energy through the breakdown of larger molecules into smaller ones

Enzymes and Cellular Metabolism

can oxidize sugars and other nutrients to harvest energy, avoids release of heat and reduces activation energy, can couple favorable reactions with unfavorable reactions

Glycolysis

overall one molecule of glucose is converted to two molecules of pyruvate with the net production of 2 ATP and 2 NADH molecules, occurs in the cytosol

3 Phases of Glycolysis

investment (2 ATP are spent to activate glucose), cleavage (glucose is split into 2 3-carbon sugars), energy generation (4 ATP molecules are generated)

Oxidation of Glyceraldehyde 3-Phosphate

favorable reaction that couples with unfavorable reaction of NADH and ATP formation to carry out glycolysis

Fermentation

oxidizes NADH in the absense of O2 to form NAD+ used for glycolysis; can lead to excretion of alcohol and CO2 or to lactate

Oxidative Metabolism

if molecular oxygen is present, aerobic organisms can further oxidize pyruvate to carbon dioxide and water

2 Steps of Oxidative Metabolism

citric acid cycle (matrix of mitochondria), electron transport chain (inner mitochondrial membrane)

Citric Acid Cycle

pyruvate is transported into mitochondrial matrix and is oxidized into Acetyl CoA and CO2; 2-carbon acetyl group is added to oxaloacetate and forms citrate; series of steps rearranges bonds and oxidizes citrate to generate CO2, reducing power (NADH/FADH2) + GTP; oxaloacetate is regenerated and the cycle restarts

Reducing Power of NADH/FADH2

energy used to reduce NAD+/FADH can be extracted to generate ATP; done by proton pumps that use NADH/FADH2 oxidation to create a proton gradient across inner mitochondrial membrane

Electron Transport Chain

NADH is oxidized to pump protons out of the mitochondrial matrix, proton gradient is then harnessed by ATP synthase (F-type pump) to make ATP

Anchoring Junctions

cell-cell or cell-matrix adhesion, cadherin or integrin proteins required for the two functions

Occluding Junctions

permeability barrier, claudin proteins required for function

Channel-Forming Junctions

intercellular passages, connexin or innexin proteins required for function

Signal-Relaying Junctions

transmit signals, anchorage and signaling proteins required for function

Intracellular Anchor Proteins

connect cytoskeletal filaments with integrins and cadherins

Cadherins

use Ca2+ and homophilic binding to connect cells together

Cells Expressing Different Types of Cadherins

sort out in vitro into clumps all composed of cells that express the same type of cadherin

Cells Expressing Different Levels of Same Cadherins

sort out in vitro into zones of cells with same levels of cadherin

Desmosomes

connect together intermediate filaments in adjacent cells to provide cells with mechanical strength

Selectins

mediate transient interactions by heterophilic binding to carbohydrates, binds to oligosaccharides on glycoproteins and glycolipids through its lectin domain

Immunoglobulin Superfamily

cell-cell adhesion molecules that have immunoglobulin (Ig)-like domains, can bind homotypically to neurons or heterotypically to integrins on white blood cells, can enable cell interactions in absense of Ca2+

Leukocytes

white blood cells that can leave the blood stream at sites of infection thanks to dynamic use of different cell-cell interactions

Tight Junctions

seal the space between adjacent membranes, seal is mediated by transmembrane proteins in bilayers of each cell, prevent diffusion of proteins in the membrane and enable transcellular support

Planar Cell Polarity

refers to the ordered arrangement of molecules in a plane, requires junctions for epithelial cells

Basal Lamina

extracellular matrix in all animal cells, organized in 3 ways: surrounds muscle cells, underlies epithelial cells, interposed between kidney glomerulus cells

Laminins

trimeric protein that can self-assemble and/or interact with many components of the extracellular matrix and with integrins to help organize the basal lamina

Integrins

connect extracellular matrix to actin cytoskeleton

Glycosaminoglycans (GAGs)

occupy large volumes of space and form hydrated gels to resist compressive forces

Collagen

major protein of the extracellular matrix, rich in lysine and proline that undergoes hydroxylation

Cell Cycle Phases

G1 phase → S phase → G2 phase → M phase

3 Major Checkpoints of the Cell Cycle

start checkpoint checks if environment is favorable and allows cell to enter cell cycle and proceed to S phase, G2/M checkpoint checks if DNA is replicated and if environment is favorable to allow cell to enter mitosis, metaphase-to-anaphase checkpoint checks if all chromosomes are attached to the spondle to trigger anaphase and proceed to cytokinesis

Control of Cell Cycle

dependent on 4 cyclically activated cyclin-dependent kinases: G1-Cdk, G1/S-Cdk, S-Cdk, and M-Cdk; control by phosphorylating different substrate proteins

Cyclical Proteolysis

cell cycle control via degrading key regultory proteins, degradation occurs through proteasome and substrates for proteasome must be ubiquitylated

Control of Proteolysis by APC/C

anaphase promoting complex or cytosome, activated in mid-mitosis and is active through the end of G1, degradation of M-cyclin occurs in the proteasome

Condensin Complex

gets duplicated chromosomes ready for separation, some of the cohesin complex is removed along the arms of the chromosome, gets ready for next step of separation of replicated DNA

Features of Model Organisms

sequenced genome, site-directed/tissue specific mutations, expression of multiple genes simultaneously across many cells, known developmental sequence

Resolution

ability to distinguish two objects close to each other

Magnification

ability to zoom in, alter field of view

Light Microscopy

standard way to view cells, allows for colored details with add-ons like fluorescence, limited by resolution

Electron Microscopy

focused beam of electrons replaces light, 200x magnification of light microscopy, only in black and white

Confocal Microscopy

scanning laser and pinhole limit detection of focal plane, creates optical section with better resolution

Flow Cytometry

analyzes cells in an aqueous stream as they pass through a laser; able to show protein interactions, reconstruct a 3D image, and observing receptor expression

Carbohydrates

polymers of sugar monomers, used as energy source and for structural support, highly polar, ex. includes monosaccharide which is uncharged in aqueous solution

Lipids

hydrophobic membrane barriers, energy sources, nonpolar molecule with hydrocarbon chain and polar COOH

Amphipathic/Amphiphilic

dislieks both water and oil, both hydrophobic and hydrophilic, property of lipids

Amino Acids

building blocks of proteins, metabolized for energy, polarity and charge depends on side group

Phospholipids

can self-assemble into bilayers; hydrophobic tails and hydrophilic heads

Monomer Linkage

form polymers under condensation reactions, specifically dehydration where water is formed and removed from the sugar

2nd Law of Thermodynamics

all processes in the universe are driven in direction that increases entropy

Why Don’t Cells Break the 2nd Law of Thermodynamics

cells are not isolated systems and can exchange energy with the environment, energy input into the cell generates order while energy from the outside of the cell is generated by processes that increase entropy, cells release heat in energy-generating processes which increases environmental entropy

Plant Cell Wall

contains cellulose (glucose polymer for tensile strength), pectin (mix of polysaccharides with cellulose for resistance to compression), and lignin (provides waterproofing)

Turgor Pressure

generation of large internal pressure in plant cell allowed by structural rigidity of wall, water flows into cell via osmosis when the intracellular environment has higher solute concentration