The insertion present at residues 102-107 creates a short ß-hairpin
near the substrate binding site.
It also contacts α-helix E2 of an adjacent
monomer, thereby increasing the contact between these particular monomers.
The second insertion is present between
α-helix E2 and ß-sheet
E at residues 141-146.
The insertion also serves to increase “subunit
interface” by contacting Phe223 on a neighboring
A notable hydrogen bonding network is present within the loop that forms.
III. Subunit Interactions
The interactions between the subunits of the ABAD/HSD10 tetramer are consistent with other SDR tetramer enzymes. Each monomer contacts the other three domains,
and the four active sites of the enzyme face toward the outside of the tetramer.
These interactions can be viewed by the three “mutually perpendicular 2-fold axes,” which relate each monomer’s position to the entire tetramer (Kissinger et al., 2004). In the ABAD/HSD10 rat homologue, type II hydroxyacyl-CoA dehydrogenase/amyloid-ß binding alcohol dehydrogenase (rHADH II/ABAD), the tetramer is formed when two dimers unite in a back-to-back manner. ABAD/HSD10 has been shown to share 88% sequence identity with rHADH II/ABAD so it may be extrapolated that the ABAD/HSD10 tetramer comes together in much the same way (Powell et al., 2000). Additionally, as noted with respect to rHADHII/ABAD, the C-terminal region of each ABAD/HSD10 monomer contacts the C-terminal region of an adjacent monomer.
IV. Substrate Binding
An unusual property of this enzyme is its ability to bind a broad range of substrates. When bound to the inhibitor AG18051
, the substrate binding region of ABAD/HSD10 takes the shape of a long and narrow cavity. This cleft consists of three main parts. Two short polypeptide chains, consisting of residues 95-99
and residues 155-168,
form one side of the substrate binding cleft. The other side of the cleft consists of the “substrate binding flap,” residues 205-217.
ABAD/HSD10’s broad substrate specificity may result from the flexibility associated with the this flap. Finally, the “floor” of the cavity is created by residues 257-261
, located at the C-terminus of the subunit. As a whole, the substrate binding region
is hydrophobic, and a series of lysine residues (Lys99, Lys104, and Lys105)
are present directly outside the cleft at the ß-hairpin of the first insertion region (residues 102-107)
In rHADH II/ABAD, the closure of the substrate binding flap (residues 205-220)
has been found to correlate directly to substrate binding and enzymatic activity. This closure may enhance enzymatic activity by blocking solvent access to the active site. In some cases this flap assumes a small helical arrangement upon substrate binding. Yet, this helix formation does not result when rHADH II/ABAD binds all possible substrates, further illustrating the broad substrate specificity of rHADH II/ABAD and ABAD/HSD10.
V. Binding of the Inhibitor, AG18051
Each monomer of the ABAD/HSD10 enzyme is complexed with NAD+
and an inhibitor.
In crystalline form, ABAD/HSD10 identically binds AG18051
at three of the four substrate binding sites of the tetramer. (When
crystallized, it appears as though the substrate binding site of one monomer
is altered and narrowed so that it is unable to bind an inhibitor, but still
). When bound, AG18051 interacts with
the cofactor, NAD+, to form a “covalent
adduct” between the N2 atom of AG18051
and the C4N atom of NAD+. Binding of AG18051
is dependent on the presence of NAD+ and does
not occur effectively when NAD+ is absent or
when NADH is present. Binding data indicates that binding of NAD+
and AG10951 is cooperative and that ABAD/HSD10
actively aids in adduct formation. Though the structure of AG18051
does not resemble that of other known ABAD/HSD10 substrates, this inhibitor
interacts closely with the substrate binding cleft
at nine specific residues.
In addition to general hydrophobic contacts, two hydrogen bonds are
formed, specifically between N1 of AG18051
of ABAD/HSD10 and between the carbonyl oxygen of AG18051
and Gln165 of ABAD/HSD10.
VI. Medical Implications:
Despite widespread interest in Alzheimer’s disease, no cure or effective
treatment has yet been developed to address this illness (Xie et al.,
2006). The onset of Alzheimer’s disease is characterized by
the build up of plaques, consisting primarily of Aß, within and surrounding
nerve cells. The presence of increased cellular levels of Aß
is thought to result in neurotoxicity, likely through a mechanism involving
the binding of ABAD/HSD10. Normally, ABAD/HSD10 plays a role in metabolic
homeostasis by helping to metabolize energy sources and to break down isoleucine
and branched-chain fatty acids (Yan and Stern, 2005). However,
when Aß is present in high concentrations, it is more likely to bind
ABAD/HSD10, resulting in the loss of function of ABAD/HSD10 due to structural
changes within the enzyme. When inhibited by Aß, ABAD/HSD10
is not able to bind and process other substrates, and the subsequent build
up of these metabolites may have adverse effects within the cell (Yan and
Stern, 2005). Recent studies also suggest that the binding of Aß
to ABAD/HSD10 induces a signal cascade that leads to apoptosis (Xie et
al., 2006). The neuronal dysfunction associated with ABAD/
Aß binding and Aß build-up can be suppressed by interrupting
the interaction between ABAD/HSD10 and Aß, a discovery which has vast
clinical possibilities. Following the release of human ABAD/HSD10’s
crystal structure and information on its binding properties (Kissinger et
al., 2004), a class of benzothiazole ureas have been shown to inhibit
ABAD/ Aß binding in an ELISA- based assay (Xie et al., 2006).
In order to develop effective AD therapeutics, more research needs to be
undertaken to determine how exactly Aß binding results in the amyloid-mediated
cell stress that is thought to cause Alzheimer’s disease and how ABAD/HSD10
functions in vivo. Because rHADH II/ABAD is very closely
related to ABAD/HSD10, one may utilize the rat model in investigating AD
and potential treatments. The molecular inhibitors of ABAD/ Aß
binding identified by Xie et al. (2006) will soon be investigated
in animal models of Alzheimer’s disease. However, because both
rHADH II/ABAD and ABAD/HSD10 play a critical role in many metabolic pathways,
difficulties may arise when trying to engineer potential therapeutics that
target this particular enzyme (Powell et al., 2000).
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Richard E. Showalter, Melwyn A. Abreo, Charles S. Agree, Stephen Margosiak,
Jerry J. Meng, Robert M. Aust, Darin Vanderpool, Bin Li, Anna Tempczyk-Russell,
and J. Ernest Villafranca. 2004. Crystal Structure of Human ABAD/HSD10 with
a Bound Inhibitor: Implications for Design of Alzheimer’s Disease
Therapeutics. J. Mol. Biol. 342: 943-952.
Powell, A. J., J. A. Read, M. J. Banfield, F. Gunn-Moore, S. D. Yan, J.
Lustbader, A. R. Stern, D. M. Stern, and R. L. Brady. 2000. Recognition
of Structurally Diverse Substrates by Type II 3-Hydroxyacyl-CoA Dehydrogenase
(HADH II)/ Amyloid-ß Binding Alcohol Dehydrogenase (ABAD). J. Mol.
Biol. 303: 311-327.
Reddy, P. Hemachandra and M. Flint Beal. 2005. Are mitochondria critical in
the pathogenesis of Alzheimer’s disease? Brain Research Reviews 49:
Xie, Yuli, Shixian Deng, Zhenzhang Chen, Shidu Yan, and Donald W. Landry. 2006.
Identification of small-molecule inhibitors of the Aß-ABAD interaction.
Bioorganic & Medicinal Chemistry Letters 16: 4657-4660.
Yan, Shi Du and David M. Stern. 2005. Mitochondrial dysfunction and Alzheimer’s
disease: role of amyloid-ß peptide alcohol dehydrogenase (ABAD). Int.
J. Exp. Path. 86: 161-171.
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