Lecture 8 Flashcards
Where is calcium concentration highest
Lowest
Outside the cell
Inside the cell (has resting (lower) and active phase (higher) )
What is calcium used as
A secondary messenger that triggers event in eukaryotic cells
What are the key things in moving calcium
PMCA (Ca2+ atpase/ ca2+ pump)
VGCC (voltage gated calcium channel)
RYR/IP3R
CLSQ (calcium sequestering)
SERCA
What is apo- CAM
Calmodulin but ca2+ Unbound form
Usually mg bound but look like apo so it’s called apo
What is serca
An atpase membrane protein in the SR/ER membrane
It pumps ca2+ into the SR/ER
What is Calsequestrin
In the SR/ER lumen
It binds a lot of ca2+ as storage in the SR/ER lumen
What is PMCA
a calcium pump in the plasma membrane
It pumps out excess ca2+ from the cell
What does calcium calmodulin do to PMCA and VGCC
Binds to their calcium binding domains to
Activate PMCA (to get ca out of cell)
Close VGCC (to make ca not get taken into cell, feedback inhibition)
VGCC is a
Heteropentamer
It’s c term inside the cell
VGCC and other ion channels are a target for
Calcium channel blocker drugs
To keep ca2+ out
How does ca cam inhibit VGCC
Thought feedback inhibition
Binds to the c term cytoplasmic part of the vgcc alpha1 subunit
This binding closes the VGCC channel
What is the inhibition of VGCC called
Calcium dependent inactivation (CDI)
Because calcium binds to cam then cam shuts off the channel
What is an isoform
Spliceoform
Diff genes
Same gene but spliced diff
How does cam bind to caV1.1 (a skeletal muscle isoform of VGCC)
The caV1.1 has a pore that allows ca2+ into the cell
can binds and pulls the two domains of the pore together
Fast inactivation
Binding of ca2+ to cam is
Sequenctial
What type of protein is calmodulin
And EF hand protein
Had 400 diff target sites (it can do many things)
What else can ca cam activate
myosin light chain kinase (MLCK)
Bind to the loop in the MLCK c term domain and pulls it off to activate it
Involved in muscle contraction
What are EF hand proteins
A protein with a EF hand motif (helix loop helix)
Connected by a small beta sheet
binds Ca in the palm of the hand
How many EF hand motifs does ca calmodulin have
4
They are organized into the n lobe and the c lobe
Will position 6 of the EF hand motif ever not be glycine
What’s always in position 1
No
Always asp in position 1
How does the EF hand bind ca
In apo state (no ca bound) the hand is closed
When ca, the hand opens and make a pocket that the substrate (whatever it’s activating) can sit in
Once ca bind to to what am happens to the structure
In apo it’s disordered, but when ca binds The structures alpha helix forms then it can bind other proteins
What do the methionine residues in cam do
How many does it have
Make cam sticky so that ca2+ binds (since hydrophobic want to be sheilded)
9 in total (4 in each lobe)
Cam is a good
Secondary messenger of proteins because it can bind in many ways
What is another enzyme that is dependent on ca cam
Ca cam dependent protein kinase 1
How does ca cam bind to Ca cam dependent protein kinase 1
Ca cam dependent protein kinase 1 has a non helical ca-cam binding domain and a helical autoinhibitory domain
This can binding domain has an exposed trp residue
This trp (trp 303) becomes an anchor for the c lobe of calmodulin so it can be pried out of the inhibitory region and be activated
What is calcineurin
A ca cam dependent phosphotase
Has alpha and beta subunits, a disordered region, and an autoinhibitory peptide
The active site of the A subunit has fe3+ and zn2+
This autoinhibitory peptide is pried off by cam, activating the phosphatase
What is in the active site of calcineurin
Fe3+ and Zn2+
What are C2 domains
Ca2+ dependent lipid binding domains
Involved in lipid binding
Conserved domains that are attached to the membrane
How is PKC (protein kinase c) activated
A hormone binds to the GPCR (ex epinephrine)
Signalling pathways make PLC (phospholipase c) cleave PIP2
IP3 (inositol triphosohate) and DAG get formed from PIP2 (they are secondary messengers)
DAG binds to PKC, PKC active only if ca is also bound
IP3 binds to the IP3 receptor (IP3R) in the ER/SR and pumps ca into the cytosol
Slide 23 know structur of IP3
Okay
Role of IP3r
In the ER/SR lumen
Release ca into cytosol
How is PKC regulated
PKC in the cytosol has a autoinhibitory region that needs to be cleaved
PKC has a C2 domain which binds ca2+ and recruits PKC to the membrane
Not active yet, so C1A and C1B binds to DAGS which cleaves off the inhibitory domain
What kind of substrates does PKC phosphorylate
Ones that are near the membrane
How do you make sure that a kinase targets one substrate and not another
Change the specificity of the c2 domain to be bound to a different lipid (so not phosserine)
Changes where PKC binds and also what substrates that surround it, this changes it’s substrates
