Download Enzymes

Important terms of enzymology
Course 2 / 210
Vladimíra Kvasnicová
Biochemical reactions are catalyzed by
enzymes:
Enzymes are commonly called according to the:
a) type of the chemical reaction
b) type of the substrate
The figure is found at: http://fig.cox.miami.edu/~cmallery/255/255enz/enzymology.htm (December 2006)
ENZYMES
Laboratory order form from http://spch.cz/kliniky/kbi/laboratorni_prirucka/zadanka_biochemie.pdf (December 2006)
„ Cardiac enzymes“
The figure is adopted from the book: Devlin, T. M. (editor): Textbook of Biochemistry with Clinical
Correlations, 4th ed. Wiley-Liss, Inc., New York, 1997. ISBN 0-471-15451-2
abbreviations of enzymes
• used in a medicine
• e.g. LD, ALT, ALP
old trivial names
• no relationship to the catalyzed reaction
• suffixed by -in (pepsin, trypsin)
• used for long time known enzymes
Enzyme nomenclature
EC nomenclature http://www.chem.qmul.ac.uk/iubmb/enzyme/
* each enzyme is classified by EC number
(Enzyme Commission of IUBMB) – 6 classes:
•
•
•
•
•
•
EC 1.x.x.x
EC 2.x.x.x
EC 3.x.x.x
EC 4.x.x.x
EC 5.x.x.x
EC 6.x.x.x
oxidoreductases
transferases
hydrolases
lyases
isomerases
ligases (synthetases)
→ classification by a reaction catalyzed by the enzyme
example:
IUBMB Enzyme Nomenclature
EC 2.7.1.1
Accepted name: hexokinase
Reaction: ATP + D-hexose = ADP + D-hexose 6-phosphate
Other name(s): hexokinase type IV glucokinase; hexokinase D;
hexokinase type IV; hexokinase (phosphorylating); ATP-dependent
hexokinase; glucose ATP phosphotransferase
Systematic name: ATP:D-hexose 6-phosphotransferase
Comments: D-Glucose, D-mannose, D-fructose, sorbitol and Dglucosamine can act as acceptors; ITP and dATP can act as donors.
The liver isoenzyme has sometimes been called glucokinase.
The reference is found at http://www.chem.qmul.ac.uk/iubmb/enzyme/EC2/7/1/2.html
systematic names
• are made according to special rules, they specify
a reaction catalyzed by the enzyme
example:
ATP : D-glucose phosphotransferase (EC 2.7.1.2)
→ transfers (2) phosphate (7) to an alcohol group (1)
ATP + D-Glc → ADP + D-Glc-6-phosphate
(Glc-6-P)
EC 2.7.1.2. = glucokinase = accepted name
oxidation / reduction (ions, organic compounds)
enzymes: oxidoreductases (EC 1.x.x.x)
distinguish:
dissociation / pH / acidity / H+ / H30+
dehydrogenation / H / H2
•
„redox equivalents“
FADH2
NADH+H+
NADPH+H+
FAD (oxidized)
FADH2 (reduced)
http://biochem.siuc.edu/web_lessons/bmb_vit.htm
http://dolly.biochem.arizona.edu/Bioc462b_Honors_Spring_2009/kyang/whatisnad2.html
common names (= accepted names) *
• simple, commonly used in practice
• very important!
1. oxidoreductases:
Aox + Bred → Ared + Box
* dehydrogenase (H- or H)
* reductase
* oxidase
* peroxidase (various peroxides)
* oxygenase (O2)
* hydroxylase (= monoxygenase; -OH)
* desaturase (-CH2CH2- → -CH=CH-)
enzymes transferring groups = transferases
(EC 2.x.x.x)
→ transfer of –NH2, phosphate, acyl, C1-fragments,...
distinguish:
acyl of an acid
anion of an acid
2. transferases:
A-x + B → A + B-x
* grouptransferase (e.g. aminotransferase)
* kinase (= phosphotransferase)
* phosphorylase
* transketolase
* transaldolase
enzymes catalyzing hydrolysis =
•
reactions:
3. hydrolases:
hydrolases (EC 3.x.x.x)
condensation / hydrolysis
A-B + H2O → A-H + B-OH
„substrate“-ase
* peptidase, glycosidase, lipase, nuclease
* esterase
(R1-CO-O-R2)
(phosphate-O-R) → Pi !!!
* phosphatase
* phosphodiesterase
(R1-O-phosphate-O-R2)
enzymes catalyzing introduction or releasing of a small compound
=
lyases (EC 4.x.x.x)
- addition or elimination of water = hydration / dehydration
- removing of CO2 = decarboxylation (not carboxylation!)
4. lyases:
A-x ↔ B + x
* decarboxylase (→ CO2)
* dehydratase
(→ H2O)
* hydratase
(-CH=CH- + H2O → -CH(OH)CH2-)
* (synthase)
enzymes catalyzing isomerization
= isomerases (EC 5.x.x.x)
isomers = chemicals having the same molecular
formula but differing in their structures
e.g. C6H12O6
5. isomerases:
glucose / fructose
A → iso-A
* epimerase (monosacharide → its epimer)
* mutase (rearangement of a phosphate group)
enzymes catalyzing synthetic reaction which needs energy from an
energy rich compound =
6. ligases:
ligases (EC 6.x.x.x)
A + B + ATP → A-B + ADP + Pi
* carboxylase
* (synthetase)
examples:
• pyruvate carboxylase
• glutamine synthetase = glutamate-ammonia ligase
Add class to which each enzyme belongs
AST
ALT
GMT
ALP
ACP
AMS
LPS
CK
CHE
LD
aspartate aminotransferase
alanine aminotransferase
gamma-glutamyl transpeptidase
alkaline phosphatase
acid phosphatase
α-amylase
lipase
creatine kinase
cholinesterase
lactate dehydrogenase
Cofactors of
oxidoreductases:
NAD+
nicotinamide adenine dinucleotide
NADP+
nicotinamide aden. dinucl. phosphate
(precursor: niacin = nicotinic acid)
HFAD
FMN
flavin adenine dinucleotide
flavin mononucleotide
2H
(precurzor: riboflavin = vitamin B2)
heme
Fe3+ + e- ↔ Fe2+
⇒
e-
transferases:
ATP
adenosine triphosphate / phosphate
GTP
guanosine triphosphate / phosphate
TDP
thiamine diphosphate / C-fragment
(prekurzor: thiamine = vitamin B1)
PALP
pyridoxal phosphate
/ -NH2
(prekurzor: pyridoxine = vitamin B6)
THF
tetrahydrofolate
/ C1-fragment
(prekurzor: folic acid)
CoA
coenzyme A (HS-Co-A)
/ acyl
PAPS
phosphoadenosine phosphosulfate / sulfate
3´-phosphoadenosine-5´-phosphosulfate (PAPS)
transfers sulfate group to a substrate (sulfatation)
http://web.indstate.edu/thcme/mwking/amino-acid-metabolism.html (Jan 2007)
Coenzyme A
= CoA-SH
http://lxyang.myweb.uga.edu/bcmb8010/pic/NAD+.gif and
http://oregonstate.edu/instruct/bb450/stryer/ch14/Slide26.jpg (Jan 2008)
Derivates of tetrahydrofolate
http://www.dentistry.leeds.ac.uk/biochem/postgrad/thftypes.gif (Jan 2008)
lyases:
PALP
ligases:
ATP
biotin
pyridoxal phosphate (decarboxylases)
adenosine triphosphate
→ acyl-CoA-synthetases
→ aminoacyl-tRNA-synthetases
= vitamin H (carboxylases)
Enzymes
•
•
•
•
•
•
lower an energy of activation (EA)
reduce the time to reach the reaction equilibrium
are not consumed or changed by the reaction
help the reaction proceed under a body´s T, p and pH
are specific
can be regulated
• don´t change the ∆G of the reaction
• don´t change the equilibrium position of the reaction
self study
The figure is adopted from the book: Devlin, T. M. (editor): Textbook of Biochemistry with Clinical
Correlations, 4th ed. Wiley-Liss, Inc., New York, 1997. ISBN 0-471-15451-2
Each enzyme has
temperature optimum
pH optimum
affinity to its substrate
The figure is found at: http://stallion.abac.peachnet.edu/sm/kmccrae/BIOL2050/Ch1-13/JpegArt113/05jpeg/05_jpeg_HTML/index.htm (December 2006)
Some enzymes are produced as precursors
(= PROENZYMES or ZYMOGENS)
The figure is found at: http://wine1.sb.fsu.edu/bch4053/Lecture26/zymogen.jpg (December 2006)
or must be activated to be active (e.g. by phosphorylation):
The figure is found at: http://fig.cox.miami.edu/~cmallery/150/memb/c11x11enzyme-cascade.jpg
(December 2006)
Isoenzymes (isozymes) are enzymes which
catalyze the same reaction but differ in their
primary structure and phyzico chemical properties
Isoenzymes are
• produced by different genes (= true isozymes)
• or produced by different posttranslational
modification (= isoforms)
• found in different compartments of a cell
• found in different tissues of an organism
• can be oligomers of various subunits (monomers)
example: 5 isozymes (various monomer ratio)
The figure is found at: http://wine1.sb.fsu.edu/bch4053/Lecture26/isozymes.jpg (December 2006)
separate enzymes of a mtb pathway
multienzyme complexes
This is Figure 17.6 from Garrett, R.H.; Grisham, C.M. Biochemistry; Saunders: Orlando,1995; page
553, found at http://www.uwsp.edu/chemistry/tzamis/enzyme_complex.html (December 2006)
example: 2-oxoacid dehydrogenase multienzyme complex
The figure is found at: http://faculty.uca.edu/~johnc/pdhrxns.gif (December 2006)
Allosteric enzyme: a) monomeric, b) oligomeric
The figure is adopted from the book: Devlin, T. M. (editor): Textbook of Biochemistry with Clinical
Correlations, 4th ed. Wiley-Liss, Inc., New York, 1997. ISBN 0-471-15451-2
Allosteric enzyme in T and R conformations: modulators shift the equilibrium
inhibitors
have a greater
affinity for
T-state
activators
and
substrates
have a greater
affinity for
R-state
The figure is adopted from the book: Devlin, T. M. (editor): Textbook of Biochemistry with Clinical
Correlations, 4th ed. Wiley-Liss, Inc., New York, 1997. ISBN 0-471-15451-2
Determination of enzyme activity for
diagnostic purposes
most often blood is investigated (serum, plasma)
→ evaluation of presence and seriousness of a
tissue damage
units: µkat/L (= catalytical concentration of enzyme)
kat = katal
1 katal = 1 mole of a substrate transformed per 1 sec
1 µkat = 10-6 kat
Enzymes found in plasma:
a) plasma-specific enzymes (e.g. clotting factors)
b) secretory enzymes (e.g. digestive enzymes)
c) cellular enzymes
Important knowledge:
1) intracellular localization of enzymes
2) organ and tissue distribution of enzymes
3) sources of enzymes found in plasma
4) way of enzyme elimination from blood
Enzyme kinetics
• activity, units
1 katal = 1 mole of a substrate transformed per 1 sec
1 IU = 1 µmole of a substrate transformed per 1 minute
1 katal
1 katal
=
1 mole
=
106 µmole
= 60 x 106 µmole
= 6 x 107 IU
/
/
/
1 sec
1 sec
1 min (= 60 sec)
The activity is related to
a constant concentration of an enzyme:
[E] = constant
The figure is found at: http://www.steve.gb.com/science/enzymes.html (December 2006)
! REMEMBER !
The figure is found at: http://fig.cox.miami.edu/~cmallery/255/255enz/gk3x15.gif (December 2006)
Michaelis-Menten kinetics
• the curve can be described by the equation:
The figure is found at: http://www.steve.gb.com/science/enzymes.html (December 2006)
Km describes an affinity of the enzyme to
its substrate
! indirect proportionality!
The figure is found at: http://fig.cox.miami.edu/~cmallery/255/255enz/gk3x15.gif (December 2006)
linearization of the curve:
y
=
kx
+
q
The figure is found at: http://www.steve.gb.com/science/enzymes.html (December 2006)
The figure is found at: http://fig.cox.miami.edu/~cmallery/255/255enz/gk3x15.gif (December 2006)
Inhibition of enzymes
The figure is found at: http://stallion.abac.peachnet.edu/sm/kmccrae/BIOL2050/Ch1-13/JpegArt113/05jpeg/05_jpeg_HTML/index.htm (December 2006)
1) Competitive inhibition
• inhibitor resembles
substrate
• it is bound to an active
site but not converted by
the enzyme
• increases Km (↓afinity of
enzyme to its S)
• if concentration of a
substrate is increased the
inhibition is decreased
• the inhibition is reversible
The figure is found at: http://www.steve.gb.com/science/enzymes.html (December 2006)
2) Noncompetitive inhibition
• inhibitor binds at a site other
than the substrate-binding
site
• inhibition is not reversed by
increasing
concentration of substrate
(no Km change)
• Vmax is decreased (it is related
to decreasing of active enzyme
concentration)
• reversible only if the inhibitor is
not bound by covalent bond
The figure is found at: http://www.steve.gb.com/science/enzymes.html (December 2006)
Summary of the inhibition
The figure is found at: http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/E/EnzymeKinetics.html
(December 2006)
Some enzymes can be inhibited also by
an excess of their substrate
The figure is found at: http://www-biol.paisley.ac.uk/Kinetics/chapter_3/chapter3_6_1.html (December 2006)
Inhibition by drugs and poisons
a) reversible
b) irreversible
→ inhibitor is bound covalently into the active
site of enzyme
Inhibition as a regulation of metabolic
pathways:
inhibition by products or intermediates:
a) feedback regulation
b) cross-regulation
c) feedforword regulation
inhibition by
d) reversible covalent modification
(e.g. phosphorylation / dephosphorylation)
Reversible covalent
modification:
A)
• phosphorylation by
a protein kinase
• dephosphorylation by
a protein phosphatase
B)
• phosphorylated enzyme
is either active or inactive
(different enzymes are
influenced differently)
The figure is found at: http://stallion.abac.peachnet.edu/sm/kmccrae/BIOL2050/Ch1-13/JpegArt113/05jpeg/05_jpeg_HTML/index.htm (December 2006)
Inhibition of enzymes used in the regulation is either
•
competitive
(Km is increased above substrate concentration within a cell)
or
• allosteric
(by conformational changes affecting the catalytic site)
Allosteric
regulation
• activator is
„a positive
modulator“
• inhibitor is
„a negative
modulator“
! the curve of allosteric enzymes is sigmoidal not hyperbolic !
The figure is found at: http://www-biol.paisley.ac.uk/Kinetics/Chapter_5/chapter5_2_2.html (December 2006)
SUMMARY
Regulation of enzyme activity
•
•
•
•
•
•
availability of a substrate and its concentration
induction of synthesis of a regulatory enzyme
activation of enzyme precursors
covalent modification of enzymes
competitive inhibition
allosteric regulation