Transcript 9- alcohol_dehydrogenase_mechanism

Mechanism of alcohol dehydrogenase
General Information on ADH
• Alcohol Dehydrogenase belongs to the
oxidoreductase family of enzymes.
• ADH is found in high concentrations
within the human liver and kidney.
General Information on ADH
• The primary and most common role of
ADH in humans is to detoxify incoming
ethanol by converting it into acetaldehyde.
• The resulting acetaldehyde, a more toxic
molecule than ethanol, is quickly
converted into acetate and other
molecules easily utilized by the cell
• The coenzymes make up a part of the
active site, since without the coenzyme,
the enzyme will not function.
Role Of NAD
• One hydrogen is
removed from alcohol
with 2 electrons as a
hydride ion (H-) while
the other is removed
as the positive ion
(H+).
Alcohol Dehydrogenase
(Oxidation-Reduction Reaction)
Only one of the protons of EtOH is abstracted and is
added to a specific side of NAD+.
H:–
O
C NH2
H
H C O H
+
+
N
H C H
H
Ethanol
H+
R
NAD+
O
Alcohol
Dehydrogenase H
O
C
CH3
H
H
C NH2
+
+
N
R
NADH
H+
• ADH is a zinc metalloenzyme 0f broad
specificity.
• It oxidizes a wide range of aliphatic,
aromatic alcohols to their corresponding
aldehydes.
• The molecule is a
symmetrical dimer,
composed of two
identical chains of
40,000 KDa each.
• Each chain has one binding site for NAD+ but
two sites for Zn 2+.
• Only one Zn ion is directly concerned with
catalysis.
• The nicotinamide ring of the NAD is bound
close to the Zinc ion at the bottom of the
pocket.
• The 2ÓH group of ribose ring is located
between the Ser-48 OH group and
imidazole group of His-51.
Active Site Characteristics of ADH
• As mentioned earlier, each subunit of one
monomer contains one binding site for NAD+
and two binding sites for Zn2+
• Each Zinc ion is ligated directly between the side
chains of Cys-46, His-67, Cys-174 and a water
molecule which is hydrogen bonded to Ser-48.
• Between the two binding sites where the zinc is
located, there are two clefts. One which binds
NAD+ and one which binds the substrate
(ethanol)
• Zinc bound to Cys-46, His-67, Cys-174, and Ser-48 (Blue) and
the coenzyme NAD+ (purple) attached to His-51 (yellow) The
eight zinc molecules are in red. The four zincs seen easily are
not directly involved in the proton transfer chain.
But we Must understand More!
• The two active sites are in clefts between the
coenzyme binding core and the catalytic
domains
• Ethanol binds to the hydrophobic core lined by
nine amino acids, which surround the substrate
• After binding NAD+, the 100 rotation makes the
protein go from its apo "open" form to the halo
"closed". This narrows the cleft, brings the
substrate binding site closer and excludes water
from the active site which is vital for the activity
of ADH
• The hydrophobic pocket:- Leu-57, Phe-93, Leu-116, Phe-110, Phe140, Leu-141, Val-294, Pro-295 and Ile-318 (red). Zinc (orange), Cys174 (purple), Cys-46 (yellow) and His-67 (green) Cxf (in this case) in
blue and oxygen involved in the dehydrogenation reaction shown
in white
Conformation Change at the Active Site
• Binding of NAD induce the release of Zn2+
water.
• Ionized alcohol displaced the Zn bound water.
• Ser 48 helps the release of hdrogen proton from
alcohol to NAD+