Upload
beck
View
50
Download
0
Tags:
Embed Size (px)
DESCRIPTION
ATP Synthase. Lecture # 14. What is ATP Synthase?. ATP (adenosine triphosphate) – the molecule of energy for cells, drives many biochemical reactions like muscle contraction, DNA and protein synthesis, etc ATP Synthase- enzyme that catalyses ATP synthesis and hydrolysis. - PowerPoint PPT Presentation
Citation preview
ATP SynthaseATP SynthaseLecture # 14Lecture # 14
What is ATP Synthase?What is ATP Synthase? ATP (adenosine triphosphate) – the molecule of ATP (adenosine triphosphate) – the molecule of
energy for cells, drives many biochemical energy for cells, drives many biochemical reactions like muscle contraction, DNA and reactions like muscle contraction, DNA and protein synthesis, etcprotein synthesis, etc
ATP Synthase- enzyme that catalyses ATP ATP Synthase- enzyme that catalyses ATP synthesis and hydrolysissynthesis and hydrolysis
http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/A/ATP.html
Where is it found?Where is it found?
A membrane enzyme:A membrane enzyme: Found in:Found in:
bacterial plasma bacterial plasma membranesmembranes
The thylacoid membrane in The thylacoid membrane in chloroplastschloroplasts
The inner mitochondrial The inner mitochondrial membrane of eukaryotic membrane of eukaryotic cellscells
http://www.biologie.uni-osnabrueck.de/biophysik/Feniouk/images/Rb_capsulatus.jpg
Parts of ATP SynthaseParts of ATP Synthase
Consists of two domains FConsists of two domains F00 and F and F11
http://www.sigmaaldrich.com/Area_of_Interest/Biochemicals/Enzyme_Explorer/Key_Resources/Metabolic_Pathways/ATP_Synthase.html
The DomainsThe Domains Hydrophobic FHydrophobic F0 0 domain sits in the membrane - domain sits in the membrane -
performs proton translocation performs proton translocation Hydrophillic FHydrophillic F11 portion protrudes from portion protrudes from
membrane - performs ATP membrane - performs ATP synthesis/hydrolysis synthesis/hydrolysis 3 alternating alpha 3 alternating alpha and beta subunits and beta subunits
http://nobelprize.org/chemistry/laureates/1997/illpres/boyer-walker.html
What does it do?What does it do?
It makes ATP from ADP and inorganic It makes ATP from ADP and inorganic phosphate (Pphosphate (Pii) )
ADP + PADP + Pi i + ATP+ ATP
The overall equation is: The overall equation is:
ADPADP3-3- + HPO + HPO442-2- + H + H++ + nH + nH++
outside membrane(+ charge)outside membrane(+ charge)
ATPATP4-4- + H + H22O + nHO + nH++inside membrane (- charge)inside membrane (- charge)
energy from protons diffusing energy from protons diffusing across membrane with the gradientacross membrane with the gradient
How does it function?How does it function?
First a proton gradient is First a proton gradient is establishedestablished Protons collect on one side of Protons collect on one side of
the membranethe membrane
Then the protons flow Then the protons flow through a channel in the through a channel in the enzyme causing the protein enzyme causing the protein subunits to rotatesubunits to rotate
http://www.sp.uconn.edu/~terry/images/anim/ATPmito.html
The “stalk” rotates in 120° The “stalk” rotates in 120° incrementsincrements
causes the units in the Fcauses the units in the F1 1
domain to contract and domain to contract and expandexpand
The structural changes The structural changes facilitate the binding of facilitate the binding of ADP and PADP and Pi i to make ATPto make ATP
Each subunit goes through 3 Each subunit goes through 3 stagesstages Open State – releases any Open State – releases any
ATPATP Loose State – ADP and PLoose State – ADP and Pii
molecules enter the subunitmolecules enter the subunit Tight State – the subunit Tight State – the subunit
contracts to bind molecules contracts to bind molecules and make ATP and make ATP
AnimationsAnimations
http://www.stolaf.edu/people/giannini/flashanimat/metabolism/atpsyn1.swf
Protons cross membrane through the ATP Protons cross membrane through the ATP synthase enzymesynthase enzyme
http://www.stolaf.edu/people/giannini/flashanimat/metabolism/atpsyn2.swf
Rotary motion of ATP synthase powers Rotary motion of ATP synthase powers the synthesis of ATP the synthesis of ATP
Interesting FactsInteresting Facts Contains 22722 atoms Contains 22722 atoms 23211 bonds connected as 2987 23211 bonds connected as 2987
amino acid groupsamino acid groups 120 helix units and 94 sheet units 120 helix units and 94 sheet units Generates over 100 kg of ATP Generates over 100 kg of ATP
daily (in humans) daily (in humans) One of the oldest enzymes-One of the oldest enzymes-
appeared earlier then appeared earlier then photosynthetic or respiratory photosynthetic or respiratory enzymesenzymes
Smallest rotary machine known Smallest rotary machine known
Picture http://webct.uga.edu/public/FRES1010CG/BCMB401098ATPSyn2.gif/Picture http://webct.uga.edu/public/FRES1010CG/BCMB401098ATPSyn2.gif/
Our ModelOur Model
Enzyme Activity and Enzyme Activity and unitsunits
Enzyme unitsEnzyme units
The enzyme unit (U) is a unit for the amount of a particular enzyme used for its activity.
“One U is the amount of the enzyme that catalyzes 1 micro mole of substrate per minute”.
The conditions also have to be specified: one usually takes as STP
Amounts of enzymes can either be expressed as molar amounts, as with any other chemical, or measured in terms of activity, in enzyme units.
Enzyme ActivityEnzyme Activity
Unit of enzyme activity:Enzyme activity = moles of substrate converted per unit time = rate × reaction volumeUsed to measure total units of activity in a given volume of solution.Specific activity:It is the amount of product formed by an enzyme in a given amount of time under given conditions per mg of total protein. = rate × reaction volume / mass of total protein.
Molecular activity: Used to compare activities of different enzymes.
Enzyme ActivityEnzyme Activity
Classical units:
Unit of enzyme activity:mol substrate transformed/min = unit
Specific activity:mol substrate/min-mg E = unit/mg E
Molecular activity:mol substrate/min- mol E = units/mol E
Enzyme ActivityEnzyme Activity
New international units:
Unit of enzyme activity:mol substrate/sec = katal
Specific activity:mol substrate/sec-kg E = katal/kg E
Molecular activity:mol substrate/sec-mol E = katal/mol E
Example 1Example 1
The rate of an enzyme catalyzed reaction is 35 μmol/min at [S] = 10-4 M, (KM = 2 x 10-5).Calculate the velocity at [S] = 2 x 10-6 M.
Work the problem.
Example 1 AnswerExample 1 AnswerThe rate of an enzyme catalyzed reaction is 35 μmol/min at [S] = 10-4 M, (KM = 2 x 10-5).Calculate the velocity at [S] = 2 x 10-6 M.
First calculate VM using the Michaelis-Menton eqn: VM [S] VM (10-4) VM (10-4)v = -----------, so: 35 = ------------------ = -------------- KM + [S] 2 x 10-5 + 10-4 1.2 x 10-4
VM = 1.2(35) = 42 mol/min; then calculate v:
42 (2 x 10-6) 84 x 10-6 v = ------------------------ = ------------ = 3.8 mol/min
2 x 10-5 + 2 x 10-6 22 x 10-6
Example 2Example 2
An enzyme (1.84 μgm, MW = 36800) catalyzes a reaction in presence of excess substrate at a rate of 4.2 μmol substrate/min. What is the TON in min-1 ? What is the TON in sec-1 ?
Work the problem.
Example 2 AnswerExample 2 Answer
An enzyme (1.84 μgm, MW = 36800) catalyzes a reaction in presence of excess substrate at a rate of 4.2 μmol substrate/min. What is the TON ?
1.84 μgm μ mol E = ------------------------- = 5 x 10-5 μmol E
36800 μgm/ μmol
4.2 μmol/min TON = ------------------ = 84000 min-1
5 x 10-5 μmol
Example 2 AnswerExample 2 Answer
What is the value of this TON (84000 min-1) in units of sec-1 ?
84000 min-1 1 sec-1
TON E = ------------------ x ---------- = 1400 sec-1
60 min-1
Example 3Example 3
Ten micrograms of carbonic anhydrase (MW = 30000) in the presence of excess substrate exhibits a reaction rate of 6.82 x 103 μmol/min.At [S] = 0.012 M the rate is 3.41 x 103 μmol/min.
a. What is Vm ? b. What is KM ?
Work these.
Example 3Example 3
a. The rate in presence of excess substrate is Vmaxso:
Vmax = 6.82 x 103 μmol/min.
b. At [S] = 0.012 M the rate is 3.41 x 103 μmol/min which is ½ Vmax so:
KM = 0.012 M. This may also be determined using the
Michaelis-Menton equation.