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PDBsum entry 1dbi
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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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Calcium-Mediated thermostability in the subtilisin superfamily: the crystal structure of bacillus ak.1 protease at 1.8 a resolution.
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Authors
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C.A.Smith,
H.S.Toogood,
H.M.Baker,
R.M.Daniel,
E.N.Baker.
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Ref.
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J Mol Biol, 1999,
294,
1027-1040.
[DOI no: ]
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PubMed id
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Abstract
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Proteins of the subtilisin superfamily (subtilases) are widely distributed
through many living species, where they perform a variety of processing
functions. They are also used extensively in industry. In many of these enzymes,
bound calcium ions play a key role in protecting against autolysis and thermal
denaturation. We have determined the crystal structure of a highly thermostable
protease from Bacillus sp. Ak.1 that is strongly stabilized by calcium. The
crystal structure, determined at 1.8 A resolution (R=0. 182, Rfree=0.247),
reveals the presence of four bound cations, three Ca(2+) and one Na(+). Two of
the Ca(2+) binding sites, Ca-1 and Ca-2, correspond to sites also found in
thermitase and the mesophilic subtilisins. The third calcium ion, however, is at
a novel site that is created by two key amino acid substitutions near Ca-1, and
has not been observed in any other subtilase. This site, acting cooperatively
with Ca-1, appears to give substantially enhanced thermostability, compared with
thermitase. Comparisons with the mesophilic subtilisins also point to the
importance of aromatic clusters, reduced hydrophobic surface and constrained N
and C termini in enhancing the thermostability of thermitase and Ak.1 protease.
The Ak.1 protease also contains an unusual Cys-X-Cys disulfide bridge that
modifies the active site cleft geometry.
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Figure 4.
Figure 4. Schematic represen-
tation of the topology of Ak.1 pro-
tease. Helices are identified by
circles and b-strands by triangles.
The location of the active site is
indicated with grey shading, with
the location of bound substrate
shown by a molecule with broken
lines.
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Figure 6.
Figure 6. Stereo diagrams showing the Ca
2+
binding sites Ca-1 and Ca-2. (a) Site Ca-1, shown with the equivalent
sites for AkP, thermitase and subtilisin BPN0 superimposed; AkP is shown in blue, thermitase in red and subtilisin
BPN0 in green. (b) Site Ca-2, shown with the equivalent sites for AkP (blue, with cyan spheres for water molecules)
and thermitase (red, with pink spheres for water molecules) superimposed. In both Figures residues are labelled as
for AkP.
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The above figures are
reprinted
by permission from Elsevier:
J Mol Biol
(1999,
294,
1027-1040)
copyright 1999.
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