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The binding of NAD to liver alcohol dehydrogenase has been studied in four
different ternary complexes by using crystallographic methods. These complexes
crystallize isomorphously in a triclinic crystal form which contains the whole
dimer of the enzyme in the asymmetric unit. This form of the enzyme has been
refined at 2.9-A resolution to a crystallographic R factor of 0.22. NAD binds in
essentially the same way in these complexes. The binding site is located at the
central part of the coenzyme binding domain. The adenine ring binds with
hydrophobic interactions between two isoleucine side chains. Both ribose rings
have 2E(C2'-endo) puckering, and each ribose makes three hydrogen bonds to the
enzyme. The pyrophosphate bridge has hydrogen bonds to the side chains of
arginine-47 and -369 and to main chain nitrogen atoms from the amino ends of two
alpha-helices. The nicotinamide ring is in van der Waals contact with the
active-site zinc atom and with the sulfur atoms of its cysteine ligands. The
carboxamide group is about 30 degrees out of the plane of the nicotinamide ring
and hydrogen bonds to main chain atoms of residues 292,317, and 319. The overall
conformation of the NAD molecule is similar to that observed for other
dehydrogenases, but differs in details. In the presence of the coenzyme, the
enzyme undergoes a large conformational change from an open to a closed form.
This conformational change has three major effects: to create favorable binding
interactions with groups of the enzyme, to enclose the coenzyme and gain binding
energy for the coenzyme by reducing the accessible surface area, and to close
off one entrance to the active site. As a comparison, ADP-ribose binding has
been studied in the open form of the enzyme. The adenosine moiety binds in a
similar way as NAD, while the rest of the molecule has different interactions.
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