BIOB10 - Cell Biology - Midterm

DNA

Stored information in double helix

Nucleotide: sugar phosphate + base (a,c,t,g)

Sugar phosphate backbone, bases connect but aren’t stuck together

DNA replication

Complimentary strand, templated polymerization, hydrolysis aids, monophosphate attaches.

Templated polymerization

Using another strand to make another strand. Using a template to make a complimentary strand.

Hydrolysis

GTP → High E bonds break → GDP + energy

Addition of water

Nucleotide

Nucleoside with extra one or more phosphate group attached (monoguanosine).

Nucleoside

The sugar and the backbone with phosphate group mentioned

Transcription

Cell needs to make sense of information. Take one strand and use it as a template to form RNA strand.

DNA → RNA

Read one at a time, RNA called transcript. Complimentary base pairings (T = U)

Translation

“What amino acid do I add?”

RNA → Protein

Read three nucleotids (1 codon) at a time. 1 codon = 1 amino acid. Ribosome does readings.

Plasma membrane

Cell is closed in. Selective barrier → how things move in and out, semi-permeable

Amphiphilic → hydrophobic tails face inward, hydrophilic head towards water

Eukaryotes

House genetic information in nucleus. Bigger genome, difference in cell structure.

Macromolecules

Like protein or nucleic acid.

Fold into energetically favourable confirmation, what makes the most sense.

Enzymes

Proteins that facilitate certain reactions. Like DNA polymerase (hydrolysis happens and it takes nucleotide away)

Nuclease

Enzyme that cuts nucleic acid

Exo: cutting from end

Endo: snipping from middle

Polypeptide chain

Multiple amino acids connected by peptide bonds.

Backbone (peptide bonds connecting), Side chain.

N-terminus front end (amino group), C-terminus back end (carboxyl group)

Hydrophobic clustering

Dropping protein into an aqueous watery environment causes proteins to move to protect parts from water.

Non-polar parts are hydrophobic.

Protein denaturing

Treat proteins with a solvent to mess up their bonds (unfold)

Protein renaturing

Remove solvent and protein folds again

Molecular chaperones

Proteins that help proteins fold properly. Folding helpers.

They do not determine shape, just help form it. Side chain interactions determine shape, exposed hydrophobic parts are meant to fold inward.

Off-pathway conformations

Exposed hydrophobic regions can result in this. Desperately going around and binding to stuff and making a mess.

Protein structures

Coiled-coil (hydrophobic face eachother), Protein complex (protein subunits join together, clump), Dimer (2, head to head arrangement. chains bind together)

Protein characteristics

Fibrous (long, coiled-coil, helix) OR globular (clump, DNA polymerase)

Can be disordered in regions (nuclear pores, gates)

Tight or weak

Specificity

Ligand binding

Referring to something that is doing bonding. Like enzymes, often first step of what an enzyme does. Fits with binding site.

Ligand

Surface contours of the molecule fit very closely to the protein. Sending communication from one molecule to another (like to change formation)

Protein binding interfaces

Surface-string, helix-helix, surface-surface (most common)

Gated transport

Pores where things can go in and out. Referring to nuclear pore complex.

Protein translocation

Proteins need to get to a destination; transmembrane proteins.

The protein uses transmembrane protein to snake through.

Vesicular transport

(main mean of protein transportation) Take a vesicle that transport from one place to another.

Signal sequences

Sorting signal. Sequences on the N-terminus or in the middle of the polypeptide chain.

Cytosolic protein

No signal sequence, is just in the cytoplasm

Co-translational translocation

Cells move proteins into the ER before completion of polypeptide synthesis.

ER sequence shows up and protein synthesis continues on ER as chain gets snaked into ER

Post-translational translocation

Finishes making protein and THEN goes to where it needs to be.

ER signal sequence

Guided to the ER by two main parts.

Signal-recognition particle (SRP) and SRP receptor.

Signal specifically on N-terminus.

Signal recognition process

SRP binds to ER signal sequence, SRP bends and block elongation factor site (pauses translation). SRP receptor binds SRP and it moves whole junction of ribosome and SRP down to ER. SRP receptor bonding. Protein elongates through protein translocator while ribosome is released.

Secretory pathway

Where stuff is going to get closer and closer to exterior of the cell

Membrane bound ribosomes

Physically on the ER, doing translation on the ER.

Free polyribosomes

Transcript with 5’ and 3’ end (read from 5’)

Ribosome keeps reading and moving down. Second and third ribosomes don’t wait till first one is done, they’ll go when there’s enough room.

Once done, ribosome disassembles into individual subunits.

Membrane-bound polyribosomes

ER signal sequence, SRP bounding to ER membrane. Continues elongating as protein snakes through. As soon as there is space, the next one attaches and binds to membrane next to first ribosome.

Sec61 complex

Core of translocator. Has a plug, when it recognizes the sequence, the plug moves aside and allows protein to enter.

Signal peptide is hydrophobic and cannot snake through

Accessory proteins

Work with translocator to help with the process of protein translocation.

BiP

BiP

Binding protein in the ER. ATP → ADP through hydrolysis. Happens in order for this to pull the protein through.

Translocon

Translocator + accessory proteins.

Targeting

(part of SRP cycle) Start-transfer sequence, signal sequence doesn’t need to stay and can be cut off.

Stop transfer.

Start-transfer sequence

It is recognized by ribosome, it opens up the translocator and enters.

Stop transfer

Get out and the little protein is embedded in the membrane

Single-pass transmembrane protein

Goes into membrane, doesn’t get cleave off, serves as an anchor. With or without stop transfer sequence. It is stuck in membrane.

Multi-pass transmembrane protein

Rest of protein snakes in, no stop transfer, just goes straight through and it’s done. Double pass if there is a stop transfer.

ER lumen protein

If they wanna be right inside the ER. One signal sequence, cleaved off by signal peptidase. Snakes right through.

ER resident proteins

Stays in the ER for whatever reason. BiP is one. Have and ER retention signal.

Protein quality control

ER must make sure protein folds correctly.

Done with Oligosaccharides and ER chaperones.

Oligosaccharides

Added to proteins. Adds three sugars to proteins (protein glycosylation), catalyzes (transferase).

ER chaperones

Proteins help proteins fold properly.

Lectins (calnexin and calreticulum)

Bind to oligosaccharides on incompletely folded proteins and retain them in ER.

Oligosaccharides processing

Adding 3 glucose to unfold protein. Glucosidase trim these off one at a time.

Retrotranslocation

The cutting process. Protein folding process doesn’t work out, it goes around the second time.

Mannosidase

Removes mannose on the core oligosaccharide tree. Not gonna get glucose added back. Mannose is removed. Protein without mannose gets destroyed.

E3 ubiquitin

enzyme. Stick onto the protein and protein translocator complex. Stamps it to be destroyed.

AAA-ATPase

Like BiP, helps pull protein out. Does this by hydrolyzing ATP. In retrotranslocation.

N-glycanase

Enzyme, still has the ubiquitin chain. It saves the chain. In retrotranslocation.

Heat-shock response

Wants more chaperones to be made, needs to go all the way back to the DNA to get that coding sequence.

Misfolding with a lot of heat → stimulates transcription of genes encoding cytosolic chaperones that help refold proteins

Unfolded protein response

Making more proteins involved in retrotranslocation, protein degredation in the cytosol, and ER chaperones

Protein translocators

Doors/gates that allow protein to pass.

Mitochondria → TOM, TIM(22/23), SAM, OXA complexes

TOM complex

Transfer proteins across the outer mitochondrial membrane. Initial recognition.

TIM complex

Transfer proteins across inner mitochondrial membrane. Continue journey down, helps bring the protein down.

SAM complex

Once protein enters TOM, this helps them fold properly in beta porin

OXA complex

Some proteins can be made in mitochondria and it does that.

Helps protein snake through, get’s cleaved in matrix. This will help it and stick it in inner membrane.

Cytosolic hsp70

(importing to mitochondrial matrix) Interacting with protein, gonna keep it unfolded. Helps that interaction. Comes off once recognized by translocator. Binds on and off to fish through.

Mitochondrial hsp60

Chaperone protein. Interacts with protein and once it comes through, this helps it fold through ATP hydrolysis.

Chloroplast

Post-translationally, translocation, ATP hydrolysis, uses TOC/TIC, added thylakoid.

Turnover number

Where the reaction rate will max out

Lysozyme

Enzyme that cut cell walls of bacteria, cell bursts. Needs to be in transition state, bound to enzyme.

Regulatory sites

Small molecules bind to these. Separate from active sites. Recognizes a regulatory molecule.

Active site

Part of enzyme that recognizes substrate, and then stuff changes when it binds with stuff.

Positive regulation

Inactive → active (both easily bind)

Negative regulation

Active → inactive (molecules don’t want the same thing)

Cooperative allosteric transitions

Binding of one inhibitor at a time, opens it up one subunit at a time. First inhibitor binds with difficulty (changes confirmation of second inhibitor in the process) and second inhibitor binds with ease.

Endocytosis

Pinching in of plasma membrane. Removes plasma membrane components and delivers them to internal compartments called endosomes.

Endocytic pathway

Leads inward from plasma membrane

Exocytosis

Secretory pathway delivers new proteins to plasma membrane or extracellular space.

Secretory pathway

Leads outward from ER towards Golgi and cell surface (or side route to lysosomes)

Coated vesicles

Cage of protein covering cytosolic surface, discard cote before fusing. Coat has 2 layers.

Clathrin-coated vesicles

Golgi to plasma membrane, 3 heavy and 3 light chains form outerlayer (triskelyons, make buds), clathrin protein component, adaptor proteins (bind to coat and trap proteins), PIPs

PIPs

Bind to AP2s and change adaptor proteins conformation. Can become phosphorylated at different regions and then bind to APs and change them. Then it is possible to bind to receptors.

They control when protein is active or inactive (no coat when inactive)

COPI

Coated transport vesicle, Golgi → ER

Arf involved

COPII

Coated transport vesicle, ER → Golgi

Sar1 involved

Membrane bending proteins

Have crescent shaped domains (BAR domains). Bind to and impose shape, help form circular bud.

Release from membrane

During uncoating of membrane, PIP that is packed into vesicles leave, this weakens adaptor protein’s bind. Adaptor proteins uncoat, as soon as they pinch off coat is useless. Uses uncoating ATPases

GEFs

GDP → GTP

Molecular switch, GTPases, activate protein, turn on

GAPs

GTP → GDP

Molecular switch, GTPases, inactivate protein, turn off.

Sar1

Inactive bound to GDP, needs to work with molecular switch. GEF takes inactive protein and activates it.

Arf

Active, bound to GTP. Works with GAPs to inactivate.

Rab-proteins

GTPases, plays a role in vesicle arriving at correct target membrane.

__-GDP is inactive

__-GEF binds and activates protein (GDP → GTP)

Tail unfolds into membrane when activated (anchor)

Rab effectors

Rab-GTP works with them, they’re tethers, catch vesicles and bring em in. Tethering proteins. Long-threadlike.

SNARE proteins

Responsible for vesicles completing membrane fusion (lipid bilayers merging to dump contents)

V: found on vesicle with 1 protein

T: found on target membrane with 3 proteins

trans SNARE complex

v- and t-SNARES wrap around to form a stable 4 helix bundle; locks the 2 membranes together for membrane fusion.

Vesicular tubular clusters

Two vesicles have their trans-SNARE complexes separated by NSF. Two vesicle’s clusters bind together and then have membrane fusion

Retrograde transport

COPII vesicles going from ER to Golgi, fusing into a large tubular cluster

Things need to go back to ER as cluster or individual (ER resident protein or COPI)

Retrieval pathway

ER resident proteins have retrieval signals if they’re sent out. BiP has KDEL sequence on C-terminal signal. Retention mechanism (ER resident protein bind to each other and form complexes too big to enter).

Golgi pathway

CGN → cis cisterna → medial cisterna → trans cisterna → TGN → lysosomes

Secretion

Leaving TGN. Destined for lysosomes (digestive enzymes), exocytosis (sent to cell membrane, straight to cell surface)

Lysosomes

Membrane enclosed organelles filled with enzymes that digest macromolecules. There is proteases and nucleases within them.

Acid Hydrolases

Enzymes in lysosomes need an acidic environment to function. Controls them from going and destroying everything. They’re physically contained and the pH isn’t suitable for them.