PDBsum entry 1b2s

Hydrolase/hydrolase inhibitor

PDB id: 1b2s

Contents

Protein chains

110 a.a. *

90 a.a. *

Ligands

SO4 ×2

Waters ×582

* Residue conservation analysis

References listed in PDB file

Key reference

Title

Structural response to mutation at a protein-Protein interface.

Authors

C.K.Vaughan, A.M.Buckle, A.R.Fersht.

Ref.

J Mol Biol, 1999, 286, 1487-1506. [DOI no: 10.1006/jmbi.1998.2559]

PubMed id

10064711

Abstract

We have crystallised three mutants of the barnase-barstar complex in which interactions across the interface have been deleted by simultaneous mutation of both residues involved in the interaction. Each mutant deletes a different type of interaction at the interface: the first complex bnHis102-->Ala-bsTyr29-->Phe (bn, barnase; bs, barstar), deletes a van der Waals packing interaction; the second complex, bnLys27-->Ala-bsThr42-->Ala, deletes a hydrogen bond; the third, bnLys27-->Ala-bsAsp35-->Ala, deletes a long-range charge-charge interaction. The contribution of each of these side-chains to the stability of the complex is known; the coupling energy between the deleted side-chains is also known. Despite each of the double mutants being significantly destabilised compared with the wild-type, the effects of mutation are local. Only small movements in the main-chain surrounding the sites of mutation and some larger movements of neighbouring side-chains are observed in the mutant complexes. The exact response to mutation is context-dependent and for the same mutant can vary depending upon the environment within the crystal. In some double mutant complexes, interfacial pockets, which are accessible to bulk solvent are formed, whereas interfacial cavities which are isolated from bulk solvent, are formed in others. In all double mutants, water molecules fill the created pockets and cavities. These water molecules mimic the deleted side-chains by occupying positions close to the non-carbon atoms of truncated side-chains and re-making many hydrogen bonds made by the truncated side-chains in the wild-type. It remains extremely difficult, however, to correlate energetic and structural responses to mutation because of unknown changes in entropy and entropy-enthalpy compensation.

Figure 3.
Figure 3. Stereo images of the site of mutation of the least-squares fit between the double mutant, bnHis102→Ala-bsTyr29→Phe, and pseudo wild-type structure. In the pseudo wild-type structure, barnase is coloured white and barstar yellow; the double mutant is coloured black. Additional water molecules, observed in the double mutant, which fill the site of mutation are also shown. (a) The AD complex. Four additional water molecules fill the created pocket. (b) The BE complex. Residues which pack against loop 2bs from a symmetry related chain, A′, are shown in black. The Figures were drawn with Bobscript [Esnouf 1997 and Kraulis 1991].

Figure 5.
Figure 5. Stereo images of the least-squares fit between the structures of the double mutant, bnLys27→Ala-bsAsp35→Ala, and the pseudo wild-type structure. Pseudo wild-type barnase is coloured white and barstar is coloured yellow; the double mutant is coloured black. Additional water molecules observed in the double mutant at the site of mutation are also shown. (a) The site of the Asp35bs→Ala mutation in the AD complex. (b) The site of the Lys27bn→Ala mutation in the AD complex. (c) The site of the Lys27bn→Ala mutation in the BE complex. The Figures were drawn with Bobscript [Esnouf 1997 and Kraulis 1991].

The above figures are reprinted by permission from Elsevier: J Mol Biol (1999, 286, 1487-1506) copyright 1999.

Secondary reference #1

Title

Protein-Protein recognition: crystal structural analysis of a barnase-Barstar complex at 2.0-A resolution.

Authors

A.M.Buckle, G.Schreiber, A.R.Fersht.

Ref.

Biochemistry, 1994, 33, 8878-8889. [DOI no: 10.1021/bi00196a004]

PubMed id

8043575

Secondary reference #2

Title

Stability and function: two constraints in the evolution of barstar and other proteins.

Authors

G.Schreiber, A.M.Buckle, A.R.Fersht.

Ref.

Structure, 1994, 2, 945-951. [DOI no: 10.1016/S0969-2126(94)00096-4]

PubMed id

7866746

Figure 1.
Figure 1. Cross-section through the barnase–barstar interface, showing some important protein–protein interactions and the residues mutated in this study. Hydrogen bonds are drawn as broken lines. This figure was drawn with the MOLSCRIPT program [32]. Figure 1. Cross-section through the barnase–barstar interface, showing some important protein–protein interactions and the residues mutated in this study. Hydrogen bonds are drawn as broken lines. This figure was drawn with the MOLSCRIPT program [[3]32].

Figure 2.
Figure 2. Representations of the structure of barstar, showing the barnase-binding surface and locations of the residues mutated in this study. Left; molecular surface of barstar colour coded according to electrostatic potential (calculated by GRASP [33]). Positively charged regions are coloured blue, negatively charged regions red. Right; backbone of barstar, drawn in the same orientation. Figure 2. Representations of the structure of barstar, showing the barnase-binding surface and locations of the residues mutated in this study. Left; molecular surface of barstar colour coded according to electrostatic potential (calculated by GRASP [[3]33]). Positively charged regions are coloured blue, negatively charged regions red. Right; backbone of barstar, drawn in the same orientation.

The above figures are reproduced from the cited reference with permission from Cell Press

Secondary reference #3

Title

Recognition between a bacterial ribonuclease, Barnase, And its natural inhibitor, Barstar.

Authors

V.Guillet, A.Lapthorn, R.W.Hartley, Y.Mauguen.

Ref.

Structure, 1993, 1, 165-176.

PubMed id

16100951

Secondary reference #4

Title

Interaction of barnase with its polypeptide inhibitor barstar studied by protein engineering.

Authors

G.Schreiber, A.R.Fersht.

Ref.

Biochemistry, 1993, 32, 5145-5150. [DOI no: 10.1021/bi00070a025]

PubMed id

8494892

Secondary reference #5

Title

Molecular structure of a new family of ribonucleases.

Authors

Y.Mauguen, R.W.Hartley, E.J.Dodson, G.G.Dodson, G.Bricogne, C.Chothia, A.Jack.

Ref.

Nature, 1982, 297, 162-164.

PubMed id

7078632