 |
PDBsum entry 1wlc
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
 |
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
Sugar binding protein
|
PDB id
|
|
|
|
1wlc
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
|
References listed in PDB file
|
|
|
Key reference
|
|
|
Title
|
|
In vitro evolutionary thermostabilization of congerin ii: a limited reproduction of natural protein evolution by artificial selection pressure.
|
|
|
Authors
|
|
C.Shionyu-Mitsuyama,
Y.Ito,
A.Konno,
Y.Miwa,
T.Ogawa,
K.Muramoto,
T.Shirai.
|
|
|
Ref.
|
|
J Mol Biol, 2005,
347,
385-397.
[DOI no: ]
|
|
|
PubMed id
|
|
|
|
|
|
Abstract
|
|
|
The thermostability of the conger eel galectin, congerin II, was improved by in
vitro evolutionary protein engineering. Two rounds of random PCR mutagenesis and
selection experiments increased the congerin II thermostability to a level
comparative to its naturally thermostable isoform, congerin I. The crystal
structures of the most thermostable double mutant, Y16S/T88I, and the related
single mutants, Y16S and T88I, were determined at 2.0 angstroms, 1.8 angstroms,
and 1.6 angstroms resolution, respectively. The exclusion of two interior water
molecules by the Thr88Ile mutation, and the relief of adjacent conformational
stress by the Tyr16Ser mutation were the major contributions to the
thermostability. These features in the congerin II mutants are similar to those
observed in congerin I. The natural evolution of congerin genes, with the
K(A)/K(S) ratio of 2.6, was accelerated under natural selection pressures. The
thermostabilizing selection pressure artificially applied to congerin II
mimicked the implied natural pressure on congerin I. The results showed that the
artificial pressure made congerin II partially reproduce the natural evolution
of congerin I.
|
|
|
|
|
|
|
|
Figure 5.
Figure 5. (a) Comparison of the congerin II wild-type
(gray) and mutant ConII-T88I (green) structures around the
mutation site T88I. The water molecules expelled by the mutation
Thr88Ile are represented by red spheres. H-bonds are shown in
yellow. (b) Comparison of the congerin II wild-type (gray) and
mutant ConII-Y16S (green) structures around the mutation site
Y16S. Bound water molecules are shown by spheres. The H-bonds in
the wild-type and mutant structures are shown in yellow and
blue, respectively. The asterisk (*)marks the position of the
loop connecting the S1-F2 strands.
|
|
Figure 6.
Figure 6. Comparison of the structures around the mutation
sites of ConII-Y16S/T88I (green) with wild-type congerin I
(blue). (a) Structures around the mutation site T88I. (b)
Structures around the mutation site Y16S. The side-chains in
wild-type congerin II are also superposed (gray).
|
|
|
|
|
|
The above figures are
reprinted
by permission from Elsevier:
J Mol Biol
(2005,
347,
385-397)
copyright 2005.
|
|
|
|
|
|
|
