Transcript What is cocaine? - Department of Chemistry

Chris Farley
Shannon Hoon
River Phoenix
What did they have
in common?
They were all cocaine
addicts and they died of
cocaine overdose!
COCAINE ESTERASE
(cocE)
The story of the ‘dope fiend’
Rhodococcus bacteria.
By: Gladys de Leon
Department of Chemistry and
Biochemistry
University of Guelph
Overview
general description of Cocaine
 Brain mechanism under influence of cocaine
 Effects and dependency
 CocE overview
 Structural and biochemical analysis of cocE
 Concluding remarks
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Structure of cocaine
What is cocaine?
C, coke, snow, nose, candy, crack
 powerful central nervous system (CNS)
stimulant
 provides intense feelings of pleasure
 Derived from leaves of South American coca
bush (erythroxylon coca and erythroxylon
novogranatense)
 Addictive and can cause death
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Erythroxylon coca
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http://cocaine.org/cokleaf.html
cultivated in Bolivia, Peru
and Ecuador
grows to a height of up to
eight feet
rich in vitamins, protein,
calcium, iron and fibre
cocaine content of the leaves
ranges from O.1% to 0.9%
History of cocaine
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Used by man since 800 A.D.
Cultivated by the Incan
1500s - Coca chewing of the South American Indians
1860s - Isolated from coca leaf by Albert Nieman
1880s - Sigmund Freud’s Uber Coca
1886 - Coca Cola a non-alcoholic medicinal tonic from
Atlanta Georgia
1890s – Cocaine containing medicine
1922 – Narcotic Drug Import and Export Act restricted
coca imports
Word from our sponsors:
Coke in the 20th Century
1 Kg of Cocaine
Tools of Trade
How does Cocaine affect the brain?
The Dopamine Hypothesis
F. I. Carroll et al, Journal of Medical Chemistry 42, 2721-36 (1999)
The brain
NA – nucleus accumbens
VT – ventral tegmental
FC – frontal cortex
CB – cerebeller
PT – putamen
CN- caudate nucleus
F. I. Carroll et al, Journal of Medical Chemistry 42, 2721-36 (1999)
What are the effects of
cocaine on the mind?
Thinking and feeling brain
 Racing thoughts
 Reward
 Felling of well being
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!! EUPHORIA !!
Negative effects:
Increased heart rate
Hyperthermia
Dilated pupils
Diaphoresis
Irritability
Decrease appetite
Sleeplessness
Depression
Over dose: respiratory arrest
Myocardial infarction
Arrhythmia
Seizures
Cocaine Dependency : Three
classical clinical characteristics that
define addiction
Psychological dependence
2. Tolerance
3. Physical withdrawal
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1. Psychological dependence
Compulsive drug seeking behavior despite
negative consequences
 Drug becomes the highest priority in the
individual’s life
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2. Tolerance
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As time goes on, more and more of the drug is
needed to produce the same high
3. Physical withdrawal
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Constellation signs and symptoms that occur
following cessation of drug use
Emergency Situation
COCAINE OVERDOSE
So what is an ideal cocaine
antidote in an emergency
setting?
High catalytic proficiency
 Lack of observable product inhibition
 Ability to hydrolyze both cocaine and
cocaethylene
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Cocaine Esterase (cocE) is an
attractive candidate for rapid
cocaine detoxification in an
emergency setting.
Overview of cocE:
Identified in the bacteria Rhodococcus sp. Strain MB1
(Rhodococcus thrives in the rhizosphere soil of the
cocaine-producing plant Erythroxylum coca)
 Cocaine degrading enzyme
 First enzyme in the metabolic pathway leading to
cocaine catabolism
 Inducible and specific for cocaine
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Hydrolysis of cocaine by cocE
D. W. Landry et al, Science, 259, 1899-1901
Cocaine Transition-State
Analogues—Abzyme Formation
Structural and Biochemical
Characterization of Cocaine
Esterase (cocE)
Structural Overview of cocE
Belongs to the / hydrolase superfamily
 574 amino acids
 ~65,000 Daltons
 3 domains – (DOM 1, DOM 2, DOM 3)
 30 % -sheet
 23 % -helix
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What is the structure of cocE?
N. A. Larsen et al., Nature Structural Biology 9, 17-21 (2002)
Domain
Structure and
Association
Domain 1: / sandwich
Residues 1-144 and
residues 241-354
 Hydrolase fold (repeating
--  motifs)
 Central -sheet is
predominantly parallel
 Contains the active site
His-287
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N. A. Larsen et al., Nature Structural Biology 9, 17-21 (2002)
REVIEW
N. A. Larsen et al., Nature Structural Biology 9, 17-21 (2002)
Domain 2: The -helical domain
95 amino acids
 Compose of 7 helices
 Inserted between 6 and
7 of Domain 1
 Five helix core – helices
H2-H6
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N. A. Larsen et al., Nature Structural Biology 9, 17-21 (2002)
REVIEW
N. A. Larsen et al., Nature Structural Biology 9, 17-21 (2002)
Domain 3: jelly-roll -barrel
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N. A. Larsen et al., Nature Structural Biology 9, 17-21 (2002)
221 amino acids
Overall fold has a jelly-rolllike topology ( i.e. HggHaemagglutinin)
Mostly -structure
-barrel-like core
Strands connected by 6 cross
over loops
Important role in the overall
tertiary structure
Active site of cocE
Located in a cleft
formed at the interface
of the three domains
 Catalytic triad – Ser
117, Asp 259 and His
287
 Oxyanion hole –
backbone amide of
Y118 and Y44
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N. A. Larsen et al., Nature Structural Biology 9, 17-21 (2002)
Proposed Mechanism for acyl
intermediate hydrolysis
Biochemical Characterization and
Structural Analysis of cocE
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cocE hydrolyzes cocaine faster than any other reported
cocaine esterase
Follows Michaelis-Menten kinetics with kcat= 7.8 s-1
and Km 640 nm.
Similar rate for cocaethylene
Validated the proposed mechanism
Reveal contributions of active site towards substrate
recognition and catalysis
J.M. Turner et al. Biochemistry, 41, 12297-12307 ( 2002)
In conclusion:
cocE enzyme itself has therapeutic potential as
an enzyme-based treatment for cocaine overdose,
furthermore, the crystal structure of the
bacterial cocaine esterase provides a basis for
further antibody engineering.
http://cocaine.org/cokehell.html
References
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W.F. Borson and T.D. Hurley, Nat Struct Biol. 9, 4-5 (2002).
N.A. Larsen et al., Nat Struct Biol. 9, 17-21 (2002).
J.M. Turner et al., Biochemistry, 41, 12297-12307 (2002).
Larsen, N. A., Zhou, B., Heine, A., Wirsching, P., Janda, K. D., and Wilson, I. A. J. Mol.
Biol. 311, 9-15 (2001)
F.I. Carroll, L.L. Howell, and M.J. Kuhar, J Med Chem. 42, 2721-2736 (1999).
L.L. Howell and K.M. Wilcox, Perspectives in Pharmacology, 298, 1- 6 (2000).
A. M. Washton and M.S. Gold. “Cocaine: A clinician’s handbook”, The Guilford Press,
New York. 1987. Pp. 73-79.
D.W. Landry et al., Science, 259, 1899- 1901 (1993).
P.H. Earley. “The Cocaine Recovery Book”, Sage Publications, London. 1991, pp.9-35.
L.M. Kamendulis et al., Jour. Phar. Exp. Ther., 279, 713-717 (1996).
M.R. Brzezinski et al., Drug Metab Dispos., 9, 1089-1096 (1997).
C. E. Mattes et al., Addict Biol., 2, 171-188 (1998).
http://www.cocaine.org/cokeleaf.html
http://www.hc-sc.gc.ca/hppb/alcohol-otherdrugs/pube/straight/stimulants.htm
http://www.a1b2c3.com/drugs/coc03.htm