PDBsum entry 1b7l

Hydrolase

PDB id: 1b7l

Contents

Protein chain

130 a.a. *

Metals

_NA

Waters ×256

* Residue conservation analysis

References listed in PDB file

Key reference

Title

Experimental verification of the 'Stability profile of mutant protein' (Spmp) data using mutant human lysozymes.

Authors

K.Takano, M.Ota, K.Ogasahara, Y.Yamagata, K.Nishikawa, K.Yutani.

Ref.

Protein Eng, 1999, 12, 663-672.

PubMed id

10469827

Abstract

The stability profile of mutant protein (SPMP) (Ota,M., Kanaya,S. and Nishikawa,K., 1995, J. Mol. Biol., 248, 733-738) estimates the changes in conformational stability due to single amino acid substitutions using a pseudo-energy potential developed for evaluating structure-sequence compatibility in the structure prediction method, the 3D-1D compatibility evaluation. Nine mutant human lysozymes expected to significantly increase in stability from SPMP were constructed, in order to experimentally verify the reliability of SPMP. The thermodynamic parameters for denaturation and crystal structures of these mutant proteins were determined. One mutant protein was stabilized as expected, compared with the wild-type protein. However, the others were not stabilized even though the structural changes were subtle, indicating that SPMP overestimates the increase in stability or underestimates negative effects due to substitution. The stability changes in the other mutant human lysozymes previously reported were also analyzed by SPMP. The correlation of the stability changes between the experiment and prediction depended on the types of substitution: there were some correlations for proline mutants and cavity-creating mutants, but no correlation for mutants related to side-chain hydrogen bonds. The present results may indicate some additional factors that should be considered in the calculation of SPMP, suggesting that SPMP can be refined further.

Secondary reference #1

Title

A general rule for the relationship between hydrophobic effect and conformational stability of a protein: stability and structure of a series of hydrophobic mutants of human lysozyme.

Authors

K.Takano, Y.Yamagata, K.Yutani.

Ref.

J Mol Biol, 1998, 280, 749-761. [DOI no: 10.1006/jmbi.1998.1906]

PubMed id

9677301

Figure 4.
Figure 4. Stereodrawing (Johnson, 1976) showing the structure in the vicinity of residue 59. (a) I59V (filled bonds) and 4SS (open bonds) structures are superimposed. (b) I59V-3SS (filled bonds) and 3SS (open bonds) structures are superimposed. Solvent water molecules are drawn as crossed circles. The broken lines indicate hydrogen bonds.

Figure 8.
Figure 8. Amino acid sequences of the wild-type (4SS), V93A, 3SS and V93A-3SS between residues 89 and 101. The substituted residues are represented in red.

The above figures are reproduced from the cited reference with permission from Elsevier

Secondary reference #2

Title

Contribution of hydrogen bonds to the conformational stability of human lysozyme: calorimetry and X-Ray analysis of six tyrosine --≫ phenylalanine mutants.

Authors

Y.Yamagata, M.Kubota, Y.Sumikawa, J.Funahashi, K.Takano, S.Fujii, K.Yutani.

Ref.

Biochemistry, 1998, 37, 9355-9362. [DOI no: 10.1021/bi980431i]

PubMed id

9649316

Secondary reference #3

Title

Contribution of the hydrophobic effect to the stability of human lysozyme: calorimetric studies and X-Ray structural analyses of the nine valine to alanine mutants.

Authors

K.Takano, Y.Yamagata, S.Fujii, K.Yutani.

Ref.

Biochemistry, 1997, 36, 688-698. [DOI no: 10.1021/bi9621829]

PubMed id

9020766

Secondary reference #4

Title

Contribution of water molecules in the interior of a protein to the conformational stability.

Authors

K.Takano, J.Funahashi, Y.Yamagata, S.Fujii, K.Yutani.

Ref.

J Mol Biol, 1997, 274, 132-142. [DOI no: 10.1006/jmbi.1997.1365]

PubMed id

9398521

Figure 2.
Figure 2. Structures in the vicinity of the residue 56 in the wild-type (a) and I56A (b) to (d), and those of the residue 59 in the wild-type (e) and I59A (f). In (c) and (d), dummy water molecules, which were estimated to be energetically favorable using the program, X-PLOR [Brunger 1992], are drawn. The dummy water molecules in (c) and (d) made hydrogen bonds with O^γ of Ser36 and an internal water molecule, respectively. The side-chain atoms of the residues 56 and 59, carbon atoms, oxygen and nitrogen atoms, interior water molecules, introduced water molecules and the dummy water molecules are represented by green, yellow, orange, blue, dark blue and purple, respectively.

Figure 3.
Figure 3. Correlation of ΔΔASA[HP] (changes in hydrophobic surface area exposed upon denaturation) with ΔΔG for the mutant proteins with empty cavities (a) and with solvated cavities (b). The mutants with solvated cavities are shown as open symbols and labeled. The mutants of the type I are represented by black filled up-triangles (with empty cavity). The black continuous line shows the linear regression of the type I mutants with empty cavity (black filled up-triangles). The mutants of the type II are represented by blue filled (with empty cavity) and open (with solvated cavity) circles. The blue broken line shows the linear regression of the type II mutants with solvated cavity (blue open circles). The mutants of the type III are represented by red filled (with empty cavity) and open (with solvated cavity) squares. The red continuous line shows the linear regression of the type III mutants with empty cavity (red filled squares). The red broken line is drawn with reference to the red continuous line. The ASA values were calculated using the procedure of [Connolly 1993].

The above figures are reproduced from the cited reference with permission from Elsevier

Secondary reference #5

Title

Contribution of hydrophobic residues to the stability of human lysozyme: calorimetric studies and X-Ray structural analysis of the five isoleucine to valine mutants.

Authors

K.Takano, K.Ogasahara, H.Kaneda, Y.Yamagata, S.Fujii, E.Kanaya, M.Kikuchi, M.Oobatake, K.Yutani.

Ref.

J Mol Biol, 1995, 254, 62-76. [DOI no: 10.1006/jmbi.1995.0599]

PubMed id

7473760

Figure 2.
Figure 2. Typical excess heat capacity curves of the mutant human lysozyme (I106V) at pH 2.70 (a), 2.92 (b), 3.04 (c), 3.10 (d), and 3.14 (e). The increments of excess heat capacity were 10 kJ/mol K.

Figure 5.
Figure 5. Stereo drawings (Johnson, 1976) showing the mutant structure in the vicinity of the mutation sites. The wild-type (open bonds) and mutant structures (filled bonds) are superimposed. (a) I23V; (b) I56V; (c) I59V; (d) I89V; and (e) I106V. Solvent water molecules are drawn as cross-circles. Broken lines indicate hydrogen bonds.

The above figures are reproduced from the cited reference with permission from Elsevier