Download Enzymes

Enzymes: A Molecular Perspective
Shuchismita Dutta, Ph.D.
Learning Objectives
• Introduction to Enzymes
• Enzyme action
• Enzyme regulation
Developed as part of the RCSB Collaborative Curriculum Development Program 2016
Learning Objectives
• Introduction to Enzymes
• Enzyme action
• Enzyme regulation
Developed as part of the RCSB Collaborative Curriculum Development Program 2016
Enzymes
Structure and Composition
• Most enzymes are proteins
• Some enzymes are RNAs
• Enzyme function (catalysis)
is facilitated by specific
amino acids or groups in the
molecule along with ions,
cofactors etc.
Functions
• Catalyzes specific reaction
• Reusable molecule
Substrate(s) + Enz Product(s) + Enz
Starch
Amylase
Salivary and Pancreatic Amylase enzymes digest starch
http://pdb101.rcsb.org/motm/74
Developed as part of the RCSB Collaborative Curriculum Development Program 2016
Enzymes Are Chemical Catalysts
• Speed up Reaction rate
• Sometimes by million fold
• Lower energy barrier
for Reaction
– Temporarily stabilizes
interaction between
substrates
– Forms Transition state
complexes
– Helps form Product(s)
http://www.ncbi.nlm.nih.gov/books/NBK9921/figure/A281/
Developed as part of the RCSB Collaborative Curriculum Development Program 2016
Amylase Digests Starch
Amylase (enzyme)
Site of substrate cleavage
Zn
Cl
Part of
starch
molecule
(substrate)
http://pdb101.rcsb.org/motm/74
Active site residues
Glu 233, Asp 197, Asp 300
Developed as part of the RCSB Collaborative Curriculum Development Program 2016
Learning Objectives
• Introduction to Enzymes
• Enzyme action
• Enzyme regulation
Developed as part of the RCSB Collaborative Curriculum Development Program 2016
Enzyme Action Models
Lock and Key
• Emil Fischer (1894)
• Active site rigid, substrate(s)
fit(s) precisely into enzyme
active site
Lock and Key
Model
Induced fit
• Daniel Koshland (1959)
• Binding of substrate(s)
changes conformations of
substrate(s) and enzyme
Induced Fit
http://www.ncbi.nlm.nih.gov/books/NBK9921/figure/A284/
Developed as part of the RCSB Collaborative Curriculum Development Program 2016
Hexokinase: Induced Fit Model
• Hexokinase (HK) transfers
a phosphate from ATP to
glucose (forms glucose 6phosphate)
• Catalyzes first step in
glycolysis
• Shaped like a clamp - big
groove in one side
• HK structure without
glucose (open); with
glucose (closed)
Hexokinase
without glucose
(open)
Hexokinase
with glucose
(closed)
http://pdb101.rcsb.org/motm/50
Developed as part of the RCSB Collaborative Curriculum Development Program 2016
Catalytic Mechanism
• Residues in catalytic site conserved in enzyme
families (e.g. Serine Proteases)
• Residues for peptide bond cleavage in all
Serine Protease enzymes
– Serine
– Histidine
– Aspartate
Trypsin
Chymotrypsin
http://pdb101.rcsb.org/motm/46
Developed as part of the RCSB Collaborative Curriculum Development Program 2016
Elastase
Substrate Specificity
Property
Trypsin
Chymotrypsin
Elastase
Enzyme
cuts after
positive amino acids
bulky amino acids
small uncharged amino
acids
Substrate
has
Lysine, Arginine
Phenylalanine,
Tryptophan
Alanine
Enzyme
specificity
pocket has
Aspartate
Serine
Valine, Threonine
Active site:
Histidine
Serine
Specificity
pocket
residues
PDB ID:2ptn
PDB ID:2cha
Developed as part of the RCSB Collaborative Curriculum Development Program 2016
PDB ID:3est
Coenzymes
• Work together with enzyme
• Small molecules, bound to
enzymes to play critical roles
• May act as carrier of specific
atoms or groups
• Are altered during enzyme
reactions
• Are recycled and may take part in
many different reactions.
– Nicotinamide adenine dinucleotide
(NAD+)
• carries electrons in oxidationreduction reactions
• can accept a hydrogen ion (H+) and
two electrons (e-) to form NADH
• NADH can donate electrons to a
second substrate, re-forming NAD+
Glyceraldehyde-3-Phosphate
Dehydrogenase active site with
bound NAD+
http://pdb101.rcsb.org/motm/5
0
Developed as part of the RCSB Collaborative Curriculum Development Program 2016
Learning Objectives
• Introduction to Enzymes
• Enzyme action
• Enzyme regulation
Developed as part of the RCSB Collaborative Curriculum Development Program 2016
Allosteric Regulation
• Binding of a small
molecule outside the
catalytic site
• Regulator binding alters
overall shape and
interactions of enzyme
• Binding may specifically
activate or inhibit
enzyme activity
http://www.ncbi.nlm.nih.gov/books/NBK9921/figure/A292
Developed as part of the RCSB Collaborative Curriculum Development Program 2016
Pyruvate Kinase: Allosteric Activation
• Pyruvate kinase makes
ATP only when needed
• Composed of four flexible
subunits arranged as a
diamond shape
• Inhibited by ATP and
amino acids
• Activated by Fructose-1,6Bisphosphate into flexed
shape.
Inactive
Enzyme
Active
Enzyme
Catlytic
Sites
Developed as part of the RCSB Collaborative Curriculum Development Program 2016
Regulatory
Sites
Enzyme Inhibition
• Why?
– To prevent excessive
function within
cells/organism
• How?
Trypsinogen: has extra
piece of protein chain to
cover active site and
keep it inactive
Pancreatic Trypsin
Inhibitor ensures that any
traces of active Trypsin
are inactivated when not
necessary
– Portion of Enzyme or
another protein blocks
access to enzyme active
site
http://pdb101.rcsb.org/motm/46
Developed as part of the RCSB Collaborative Curriculum Development Program 2016
Enzyme Inhibitors as Drugs
• HIV-1 Protease bound
to substrate peptide
• HIV Protease inhibitors
as Drugs: Substrate-like
or other molecules
bind to protease active
site  block HIV-1
protease function (viral
maturation)
Indinavir
Saquinavir
Ritonavir
Nelfinavir
http://pdb101.rcsb.org/motm/6
Developed as part of the RCSB Collaborative Curriculum Development Program 2016
Summary
• Introduction to Enzymes
– Enzymes and Catalysis
• Enzyme action
– Enzyme Action models
– Catalytic mechanisms, specificity and coenzymes
• Enzyme regulation
– Allosteric Regulation
– Inhibition and Drugs
Developed as part of the RCSB Collaborative Curriculum Development Program 2016