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PDBsum entry 1vfo
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References listed in PDB file
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Key reference
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Title
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Complex structures of thermoactinomyces vulgaris r-47 alpha-Amylase 2 with acarbose and cyclodextrins demonstrate the multiple substrate recognition mechanism.
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Authors
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A.Ohtaki,
M.Mizuno,
T.Tonozuka,
Y.Sakano,
S.Kamitori.
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Ref.
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J Biol Chem, 2004,
279,
31033-31040.
[DOI no: ]
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PubMed id
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Abstract
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Thermoactinomyces vulgaris R-47 alpha-amylase 2 (TVAII) has the unique ability
to hydrolyze cyclodextrins (CDs), with various sized cavities, as well as
starch. To understand the relationship between structure and substrate
specificity, x-ray structures of a TVAII-acarbose complex and inactive mutant
TVAII (D325N/D421N)/alpha-, beta- and gamma-CDs complexes were determined at
resolutions of 2.9, 2.9, 2.8, and 3.1 A, respectively. In all complexes, the
interactions between ligands and enzymes at subsites -1, -2, and -3 were almost
the same, but striking differences in the catalytic site structure were found at
subsites +1 and +2, where Trp(356) and Tyr(374) changed the conformation of the
side chain depending on the structure and size of the ligands. Trp(356) and
Tyr(374) are thought to be responsible for the multiple substrate-recognition
mechanism of TVAII, providing the unique substrate specificity. In the beta-CD
complex, the beta-CD maintains a regular conical structure, making it difficult
for Glu(354) to protonate the O-4 atom at the hydrolyzing site as a previously
proposed hydrolyzing mechanism of alpha-amylase. From the x-ray structures, it
is suggested that the protonation of the O-4 atom is possibly carried out via a
hydrogen atom of the inter-glucose hydrogen bond at the hydrolyzing site.
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Figure 3.
FIG. 3. Stereo views of interactions between catalytic site
residues (light gray) and acarbose (gray)(a),  -CD
(gray)(b),  -CD (gray) (c), and  -CD
(gray)(d), illustrated by the program MOLSCRIPT (23). Trp356 and
Tyr374 in the unliganded TVAII are superimposed (white). The
selected hydrogen bonds are shown with dotted lines.
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Figure 5.
FIG. 5. The proposed hydrolyzing mechanism for substrate
with a linear structure (a) and the hypothetical -CD
hydrolyzing mechanism of TVA II (this study) (b) are shown.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2004,
279,
31033-31040)
copyright 2004.
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