PDBsum entry 1xg6

Hydrolase inhibitor

PDB id: 1xg6

Contents

Protein chain

179 a.a.

Waters ×190

References listed in PDB file

Key reference

Title

Single mutation at p1 of a chymotrypsin inhibitor changes it to a trypsin inhibitor: X-Ray structural (2.15 a) and biochemical basis.

Authors

S.Khamrui, J.Dasgupta, J.K.Dattagupta, U.Sen.

Ref.

Biochim Biophys Acta, 2005, 1752, 65-72.

PubMed id

16081330

Abstract

Change in specificity, caused by the mutations at P1 site, of the serine protease inhibitors of different families is reported in the literature, but Kunitz (STI) family inhibitors are almost unexplored in this regard. In this paper, we present the crystal structure of a P1 variant of winged bean chymotrypsin inhibitor (WCI) belonging to Kunitz (STI) family, supplemented by biochemical, phylogenetic and docking studies on the mutant. A single mutation (Leu-->Arg) at P1 converted WCI to a strong inhibitor of trypsin with an association constant of 4.8x10(10) M(-1) which is comparable to other potent trypsin inhibitors of the family. The crystal structure (2.15 A) of this mutant (L65R) shows that its reactive site loop conformation deviates from that of WCI and adopts a structure similar to that of Erythrina caffra trypsin inhibitor (ETI) belonging to the same family. Mutation induced structural changes have also been propagated in a concerted manner to the neighboring conserved scaffolding residue Asn14, such that the side chain of this residue took an orientation similar to that of ETI and optimized the hydrogen bonds with the loop residues. While docking studies provide information about the accommodation of non-specific residues in the active site groove of trypsin, the basis of the directional alteration of the reactive site loop conformation has been understood through sequence analysis and related phylogenetic studies.

Secondary reference #1

Title

Refined crystal structure (2.3 a) of a double-Headed winged bean alpha-Chymotrypsin inhibitor and location of its second reactive site.

Authors

J.K.Dattagupta, A.Podder, C.Chakrabarti, U.Sen, D.Mukhopadhyay, S.K.Dutta, M.Singh.

Ref.

Proteins, 1999, 35, 321-331. [DOI no: 10.1002/(SICI)1097-0134(19990515)35:3<321::AID-PROT6>3.3.CO;2-P]

PubMed id

10328267

Figure 5.
Figure 5. The first reactive site of WCI (green) is docked at the enzyme (magenta) active site (van der Waals surface is shown). shows His57 at the center having a hydrophobic environment (partially shown for clarity in viewing).

Figure 8.
Figure 8. The canonical conformation of the second reactive site loop. The side-chain of Glu39 intrudes inside the loop and stabilizes it through hydrogen bonding.

The above figures are reproduced from the cited reference with permission from John Wiley & Sons, Inc.

Secondary reference #2

Title

The role of asn14 in the stability and conformation of the reactive-Site loop of winged bean chymotrypsin inhibitor: crystal structures of two point mutants asn14--≫lys and asn14--≫asp.

Authors

S.Ravichandran, J.Dasgupta, C.Chakrabarti, S.Ghosh, M.Singh, J.K.Dattagupta.

Ref.

Protein Eng, 2001, 14, 349-357.

PubMed id

11438758