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PDBsum entry 1ee2
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Oxidoreductase
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PDB id
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1ee2
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Contents |
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* Residue conservation analysis
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References listed in PDB file
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Key reference
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Title
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Structural basis for substrate specificity differences of horse liver alcohol dehydrogenase isozymes.
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Authors
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H.W.Adolph,
P.Zwart,
R.Meijers,
I.Hubatsch,
M.Kiefer,
V.Lamzin,
E.Cedergren-Zeppezauer.
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Ref.
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Biochemistry, 2000,
39,
12885-12897.
[DOI no: ]
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PubMed id
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Abstract
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A structure determination in combination with a kinetic study of the steroid
converting isozyme of horse liver alcohol dehydrogenase, SS-ADH, is presented.
Kinetic parameters for the substrates, 5beta-androstane-3beta,17beta-ol,
5beta-androstane-17beta-ol-3-one, ethanol, and various secondary alcohols and
the corresponding ketones are compared for the SS- and EE-isozymes which differ
by nine amino acid substitutions and one deletion. Differences in substrate
specificity and stereoselectivity are explained on the basis of individual
kinetic rate constants for the underlying ordered bi-bi mechanism. SS-ADH was
crystallized in complex with 3alpha,7alpha,12alpha-trihydroxy-5beta-cholan
-24-acid (cholic acid) and NAD(+), but microspectrophotometric analysis of
single crystals proved it to be a mixed complex containing 60-70% NAD(+) and
30-40% NADH. The crystals belong to the space group P2(1) with cell dimensions a
= 55.0 A, b = 73.2 A, c = 92.5 A, and beta = 102.5 degrees. A 98% complete data
set to 1.54-A resolution was collected at 100 K using synchrotron radiation. The
structure was solved by the molecular replacement method utilizing EE-ADH as the
search model. The major structural difference between the isozymes is a widening
of the substrate channel. The largest shifts in C(alpha) carbon positions (about
5 A) are observed in the loop region, in which a deletion of Asp115 is found in
the SS isozyme. SS-ADH easily accommodates cholic acid, whereas steroid
substrates of similar bulkiness would not fit into the EE-ADH substrate site. In
the ternary complex with NAD(+)/NADH, we find that the carboxyl group of cholic
acid ligates to the active site zinc ion, which probably contributes to the
strong binding in the ternary NAD(+) complex.
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Secondary reference #1
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Title
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Substrate specificity and stereoselectivity of horse liver alcohol dehydrogenase. Kinetic evaluation of binding and activation parameters controlling the catalytic cycles of unbranched, Acyclic secondary alcohols and ketones as substrates of the native and active-Site-Specific co(ii)-Substituted enzyme.
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Authors
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H.W.Adolph,
P.Maurer,
H.Schneider-Bernlöhr,
C.Sartorius,
M.Zeppezauer.
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Ref.
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Eur J Biochem, 1991,
201,
615-625.
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PubMed id
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Secondary reference #2
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Title
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Preparation and characterization of isozymes and isoforms of horse liver alcohol dehydrogenase.
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Authors
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I.Hubatsch,
P.Maurer,
D.Engel,
H.W.Adolph.
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Ref.
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J Chromatogr A, 1995,
711,
105-112.
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PubMed id
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Secondary reference #3
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Title
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Electrostatic effects in the kinetics of coenzyme binding to isozymes of alcohol dehydrogenase from horse liver.
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Authors
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H.W.Adolph,
M.Kiefer,
E.Cedergren-Zeppezauer.
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Ref.
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Biochemistry, 1997,
36,
8743-8754.
[DOI no: ]
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PubMed id
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