Lecutre - Protein Shit Flashcards
Concentration of oxygen is saline vs heamogolin in blood
The concentration of oxygen in saline solution is limited to ~ 0.2 mmol/L
- while the concentration of haemoglobin in blood is ~ 5 mmol/L.
Highly active tissue, e.g. exercising muscle or the brain, is limited by…
… availability of oxygen
There is strong evolutionary pressure for
Efficient oxygen transport and delivery
Active tissues use more ____ then _____ can deliver
Active tissues can use more O2 then blood can deliver
Myglobin meets challenge of needing lots of oxygen by…
… storing oxygen in muscle against Burts of high requirement
Globin
- myomolgobin is red and stored in tissues
How much myoglobin in blood
Human muscles have 0.5-0.7 mmol/L myoglobin, enough for about 7 seconds of intense activity. After this store is exhausted the tissue depends on oxygen delivery from the lungs.
Primary protein structure of myoglobin
150 amino acids
Protein secondary structure of myoglobin
Eight a-helices A-H and connecting loops (AB, BC, etc.)
Tertiary structure of myoglobin
globin fold with a hydrophobic pocket
Myoglobin quanternary structure
monomeric (a single polypeptide chain)
What does the ancient globin fold provide?
- hydrophobic pocked (see Val E11 and Phe CD1, next slide) to bind a haem group.
Haem is a…
…prosthetic group or cofactor
Structure if Haem
- includes pyrrole rings linked together in a plane
- iron has 6 coordinate bonds - four to nitrogen atoms of the Haem, one to a nitrogen atom of histidine F8 the globin, one to O2
- binding of oxygen to Fe2+ is a reverable reaction
What gives Haem its red colour
- the molecular electronic orbitals of Haem give it a red colour
Binding of oxygen to the Fe2+ is a….
.. reversible intereaction
Haem
Spectroscopy quantifies..
..dissolved molecules
Haem is rich in..
Carbon and nitrogen
- they all bind an iron atom where oxygen binds
Haem e planar?
Nah not quite - close to flat with fe in middle
HisF8 holds it in place - can also sense what’s going on thriugh bond
- side chains prevent oxygen from binding straight up and binding too strongly in an ideal postition
Which interactions break and reform dousing myoglobins function
- His E7’s interaction with oxygen
- oxygens interation with iron
What measures oxygens binding to myoglobin
Spectroscopy
How spectroscopy quantifies dissolved molecules - yappage
- higher concentration = less transmitted light = higher absorbance
- beer-lambert law converts from absorbance to concentration
- monochromator - breaks light into its diff coulours - pushes known amount of light into solution - certain amout gets absorbed
- when converting between nitrogen bound and oxygen bound - can say the haemoglobin isn’t changing - only whether its in an oxygen bound or oxygen free state
What measures binding and unbinding of Haem ogle in
Spectroscopy
Spectroscopy of globin measure oxygen binding
- shape of spectrum differs with colour and with chemical nature of solute
- protein is colourless (but has UV absorbance)
- Haem has visible absorbance (and therefore colour) that differs between bright red oxyhaemoglobin (HbO2)dull red deoxyhaemoglobin (Hb)
Haem Fe2+ is attached to globin protein by
Co-ordinate linkage to His F8
His E7 is located where and why
..on the opposite side of haem and distorts binding of gas molecules to 6th co-ordination position of haem Fe2+
- this reduces the binding affinity of oxygen to myoglobin, making it easier to release oxygen to the muscle cell
Myoglobin and haemoglobin both show ______ ______ of oxygen affinity
Myoglobin and haemoglobin both show ‘allosteric control’ of oxygen affinity
- allosteric means ‘without overlapping’
- allosteric builds on steric hindrance - the impossibility of two atoms occupying the same space
What does lactate do the myoglobins affinity for oxygen?
Lactate decreases myoglobins affinity for oxygen but does not bind where oxygen bins
- this shifts the curve right wards
- lactate build up in muscles promotes oxygen release from myoglobin, increasing O2 availability for respiration
Graph of myoglobin and haemoglobin differing in O2 binding
- myoglobin is saturated with O2 at low pO2 only releasing O2 to muscle cells when the cellular pO2 is very low
- this is shown in a hyperbolic curve
- this suits its function as a ‘back-up’ store of O2 in muscle cells
Mb + O2 = MbO2
The partial pressure (pO2) of oxygen in lungs vs resting muscle
100 Torr in lungs
20 Torr in resting muscle
Hamaglobion oxygen binding curve shape
Sigmoidal
Compared to myoglobin, hemoglobin funcitoning as an O2 transporter in blood evolved a much weaker….
… binding affinity for oxygen
The availability of O2 to cellular proteins depends on:
- the pO2 in the local environment
- the binding affinity of O2 to haemoglobin
How haemoglobins function means it has envelopes to bind O2 less tightly then myoglobin
- bind O2 in the vicinity of the lungs where the pO2 is 100Torr
- release the O2 in the vicinity of peripheral tissues where the pO2 is 20 Torr
- it can also change shape, ‘duping’ oxygen in peripheral tissues
Which binds haemoglobin more tight?
Myoglobin
Haemoglobin has evolved to be a (Structure)
Tetramer - four glob in proteins associate together non-covalently
- each globin protein contains one haem and each can bind one O2
- 1,2,3,4 O2 can bind per one Hb tetramer
Myoglobin (a monomer) acts as a
O2 store in muscle
Structure difference of deoxyhaemoglobin and oxyhemoglobin
When was the MWC concerted model proposed and who by?
Proposed in 1956 by Monod,
Wyman and Changeux
MWC, concerted model
- Subunits can be in a low-activity, tense (T) or high-activity, relaxed (R) conformation.
- All subunits must be in the same state (either T or R).
- Binding each successive substrate (S) shifts equilibrium in favour of R.
- Inhibitors stabilise the T form.
- Activators stabilise the R form.
KNF, sqential model ([prolly not in exam)
• One substrate binding induces a T → R conformational
change in only one subunit.
• This conformational change influences the neighbouring
subunits (i.e. cooperativity), changing their affinity for
substrate. Many conformations are possible.
• This explains both positive and negative cooperativity.
Similarities between haemoglobin and myoglobin
• Oxygen binds to iron of haem.
• Shift from dull to bright red allows monitoring O 2
binding.
• Affinity for oxygen is altered by molecules (e.g. lactate to
myoglobin) binding elsewhere (allosteric control).
Myoglobin vs haemoglobin - differences
• Monomer versus tetramer
• Tighter, hyperbolic versus weaker, sigmoidal binding curve
• Store in tissue versus transport molecule
Cooperatively
- requires multiple interacting subunits
- gives a sigmoidal binding curve
- shifts binding affinity (and the steep part of the binding curve) to a physiologically relavant oxygen concentration
