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75 notecards = 19 pages (4 cards per page)

Viewing:

BMB424-EX2

front 1

FUCCI

back 1

Uses Cdt1 (G1 phase = red) and Geminin → (S/G2/M phases = green)

yellow = transition

front 2

Brdu Labeling

back 2

Uses synthetic nucleoside analog of thymidine, then use antibodies against BrdU

sees where replicating DNA is

flourencent

front 3

19S cap

back 3

recognize polyubiquitin

recycle ubiquitin

unfolding

front 4

20s cap

back 4

chymotrypsin --> cut after hydrophobic

Trypsin --> cut after basic

Caspase --> cutes after acidic

front 5

Mono vs Multi vs Poly ubiquitination

back 5

Mono - attach 1 ub (trafficking)

Multi - multiple sites (endocytosis)

Poly - long chain of ub (degradation)

front 6

Impact of dwell time

back 6

The longer a protein can be associated with the regulatory subunit of the
proteosome, the more likely the protein is going to be degraded

front 7

How does rRNA contribute to translational efficiency

back 7

rRNA is most of the ribosome

catalyzes bind formation for speedy translation

anticodon bonds to only specific mRNA+tRNA

front 8

Ligand

back 8

signal

sends info from one cell to another

front 9

Receptor

back 9

protein on target cell to receive signal

front 10

Transduction

back 10

Relayer

Chain reaction to pass message along

front 11

Response

back 11

cell changing

front 12

Molecular switches (example)

back 12

switching between on + off

GTP binding (bound to GTP = ON)

front 13

Cell signal specificity

back 13

receptors unique to certain signals

different downstream proteins

front 14

Collins et al

back 14

26S Proteasome --> degradation of proteins

ubiquitination doesn't equal immediate degradation (dwell time)

front 15

Ribosome Profiling Density

back 15

More reads/higher density = more ribosome

front 16

Ribosome Profiling Positions

back 16

clustered in codon reading = active translation

peaks at start codon = initiation

front 17

Ribosome Profiling high build ups

back 17

ribosome slowing or stalling to read

front 18

Ribosome Profiling Translation efficiency

back 18

High mRNA + low ribosome = inefficient

front 19

Positive feedback

back 19

outputs make the process faster

cell division

On diagram: loops, plus signs, promoting production

front 20

Negative feedback

back 20

outputs shut down process

inhibition

blunt ends, minus signs, product shuts down production

front 21

Methylation

back 21

creates distinct regions in chromatin regulation

keeps charge

changes protein - protein interactions

front 22

Acetylation

back 22

neutralizes charge (+ histones hugging - DNA)

opens chromatin to activate genes

front 23

Glycosylation

back 23

adding sugars for folding, stability, and recognition

ID in ER

front 24

Phosphorylation

back 24

adds large - charged group

on/off switch

assembles large proteins into complexes

front 25

Cell cycle check points

back 25

G1/s - transition/start (size? environment? density? attachment?)

G2/M - mitosis (replicated? environment?)

Meta to Ana - cytokinesis (chromosomes attached? ready to split?)

front 26

Cyclin classes

back 26

G1 - high levels = expression of G1/S cyclins

G1/S - activate cdk to push cell into S phase for cell cycle entry

S - stimulate chromosome duplication

M - enter mitosis at G2/M and degrade at meta to ana

front 27

EF-Tu

back 27

EF1

escorts tRNA to A site

front 28

EF-G

back 28

EF2

uses GTP hydrolysis to provide mechanical push of ribosome forward

front 29

RF

back 29

Stop codons

front 30

Transportation into ER + sorting

back 30

entry for secretory pathway

use vesicles to go to golgi apparatus

sends to final destination

front 31

Major roles of ER

back 31

Manufacture - lipids + export proteins

Quality control - check proteins before fold

Storage - store Ca2+ for signal

front 32

ER resident chaperones

back 32

BiP - helpers - grab floppy proteins and folds correctly

Disulfide bonds - staples - keep together

front 33

ER bound protein translation

back 33

water freezing aa comes out > SRP grabs ribosomes to stop > SRP receptot docks ribosome > translocator in membrane for protein

front 34

ER sequence signals

back 34

Start transfer - opens door for threading

Stop transfer - closes door for threading

front 35

How would a protein on membrane orient based on ER signaling sequence?

back 35

Standard signal usually cut off

N term in ER with C term in cytosol (BC C is +)

front 36

Class of cell surface receptors

back 36

Ion channels

g protein using (GPCR)

enzyme coupled

front 37

Second messengers

back 37

molecules that spread signals deep into a cell after receptor activation

cAMP

front 38

Fast vs Slow signaling

back 38

Fast - exocytosis

Slow - cell cycle entry

front 39

GCPR activation (Gs vs Gi)

back 39

Gs - activates adenylyl cyclase

Gi - inhibits adenylyl cyclase

front 40

GCPR activation steps

back 40

ligand binds to GCPR > GDP kicked out > GTP let in > split into a and B > signal stops once a subunit times out > GTP turned back to GDP

front 41

How is the 26S proteasome processive?

back 41

26S proteasome is processive because it binds, unfolds, translocates, and fully degrades proteins in one continuous operation without releasing them halfway

front 42

Ubiquitination process

back 42

E1 picks up + activates Ub with ATP > E2 carries live Ub from E1 to E3 > E3 acts as a matchmaker for Ub and specific target protein

front 43

Yip et al

back 43

UBE20 binds best with Ub and NAP1L1

front 44

Formicheva et al

back 44

NF-kB family of transcription factors

noncanonical path implicated in regulating cell signaling

high density = stop cell division

front 45

Ignolia et al

back 45

Measure translation genome wide at single codon resolution

front 46

Kinase structure + activity

back 46

Small N term, large C term, active site in middle

T loop regulation (phosphorylated = moving loop and active site to be available)

front 47

Non proteolytic signaling

back 47

changing location/activity

front 48

tRNA

back 48

bridge between mRNA codon + aa

anticodon binds w tRNA

front 49

RPN 1, 10, 13

back 49

initial Ub binding

front 50

USP 6, 14 + UCH37

back 50

ub removal + 19S activation

front 51

RPN11

back 51

removing final Ub

front 52

RPT1-6

back 52

substrate binding + unfolding

front 53

RPT 2, 3, 5

back 53

20s gate open

front 54

A vs P vs E site

back 54

A - aminoacyl - charged tRNA entry

P - peptidyl - holding tRNA attached to growing polypeptide

E - exit - uncharged DNA

front 55

Lipidation

back 55

anchors protein to cell membrane since fats like to be near each other

front 56

M cdk

back 56

assembles miotic spindles

ensure sister chromatids apart

chromosome condensation

break down of nuclear envelope

front 57

Ribosome profiling procedure

back 57

freeze ribosome > lyse cells > digest unprotected RNA > isolate ribosome > convert to cRNA > sequence > align

front 58

Cyclins

back 58

cell division regulating proteins

activated when needed

front 59

PTM regulation of cdks

back 59

phosphorylates T loop

wee1 adds P but not active til cell is ready

cdc25 removes inhibiting P --> burst of energy

front 60

Proteasome check + balance

back 60

DUBs > removing Ub to recycle

Narrow 20S gate

front 61

Cell cycle phases

back 61

Interphase (G1, S, G2) - growing, synthesizing, growing

M phase - mitosis

Cytokinesis - splitting

front 62

Prophase

back 62

Chromosome condense

front 63

Prometaphase

back 63

nuclear envelope breaks

front 64

Metaphase

back 64

chromosomes line up in middle

front 65

Anaphase

back 65

sister chromatins pulled apart

front 66

Telophase

back 66

2 nucli formed

front 67

cdks

back 67

phosphorylates target

T loop can expose active site

front 68

Anaphase Promoting Complex (ACP/C)

back 68

catalyzes ub in exit of mitosis

cleans up

ends cell division and destroys proteins w cyclins

front 69

P21 and P27

back 69

P21 - safety brake - stops if DNA damaged

P27 - stop and stay - signal to stop growth in G1

front 70

cdc20 and cdh1

back 70

cdc20 - triggers ACP/C activity

cdh1 - maintains ACP/C activity after anaphase

clean up

front 71

SCF

back 71

destroys P27 so cell can move from G1 to S

target must be phosphorylated

front 72

CDK1 to active M cdk

back 72

cdk1 + M cyclin = inactive M cdk

CAK + wee1 + inactive M cdk = inactive M cdk with active P and inhibiting P

cdc25 takes inhibiting P and creates active M cdk

front 73

ACP/C...

+cdh1

+cdh20

back 73

cdh1 - keeps cyclin levels low so cell can grow and not reenter S phase

cdh20 - destroys securin + M cyclins

front 74

Sequential activation of cdks

back 74

G1 - cdk4/6 - cyclin D - kicks off mitosis

G1/S - cdk2 - cyclin E - push cell past start

S - cdk2 - cyclin A - triggers DNA replication

M - cdk1 - cyclin B - cell division

front 75

K11 vs K63 vs K48 vs K29/33

back 75

K11 -marks cyclins for rapid degradation in mitosis

K63 - DNA repair

K48 - degrade proteasome, unfolds so protein can be shredded

K29/33 - protein trafficking