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Single amino acid changes in the Bacillus thuringiensis var. israelensis δ-endotoxin affect the toxicity and expression of the protein

Single amino acid changes in the Bacillus thuringiensis var. israelensis δ-endotoxin affect the toxicity and expression of the protein
Single amino acid changes in the Bacillus thuringiensis var. israelensis δ-endotoxin affect the toxicity and expression of the protein

Site-directed mutagenesis has been used to change individual amino acids of the larvicidal 27,000 Mr, δ-endotoxin of Bacillus thuringiensis var. israelensis. Basic and acidic residues have been systematically replaced by alanine, and the resulting mutant polypeptides analysed for cytolytic and larvicidal activity, and binding to phosphatidyl choline liposomes. Replacement of residues at positions 154, 163, 164, 213 and 225 results in proteins which accumulate as inclusions in recombinant Bacillus subtilis cells similar to the wild-type, but have considerably reduced in-vitro and in-vivo toxicity. One mutant (Glu45 to Ala45) results in a protein that has reduced activity in vitro, but retains wild-type larvicidal toxicity. In addition, seven other mutations of charged residues result in proteins which form small or no inclusions in recombinant cells, despite being produced at levels similar to the wild-type in six out of seven cases. In most instances, the toxicity of these aberrantly expressed proteins is considerably less than the wild-type, although one (Lys124 to Ala124) results in a polypeptide with approximately threefold increased activity in vitro. A secondary structural model is proposed to explain these observations.

0022-2836
527-535
Ward, E. S.
b31c0877-8abe-485f-b800-244a9d3cd6cc
Ellar, D. J.
92e52b64-6600-401e-a3d0-b15cfff77199
Chilcott, C. N.
80bf4b78-c610-4c21-aad3-85f371537442
Ward, E. S.
b31c0877-8abe-485f-b800-244a9d3cd6cc
Ellar, D. J.
92e52b64-6600-401e-a3d0-b15cfff77199
Chilcott, C. N.
80bf4b78-c610-4c21-aad3-85f371537442

Ward, E. S., Ellar, D. J. and Chilcott, C. N. (1988) Single amino acid changes in the Bacillus thuringiensis var. israelensis δ-endotoxin affect the toxicity and expression of the protein. Journal of Molecular Biology, 202 (3), 527-535. (doi:10.1016/0022-2836(88)90283-5).

Record type: Article

Abstract

Site-directed mutagenesis has been used to change individual amino acids of the larvicidal 27,000 Mr, δ-endotoxin of Bacillus thuringiensis var. israelensis. Basic and acidic residues have been systematically replaced by alanine, and the resulting mutant polypeptides analysed for cytolytic and larvicidal activity, and binding to phosphatidyl choline liposomes. Replacement of residues at positions 154, 163, 164, 213 and 225 results in proteins which accumulate as inclusions in recombinant Bacillus subtilis cells similar to the wild-type, but have considerably reduced in-vitro and in-vivo toxicity. One mutant (Glu45 to Ala45) results in a protein that has reduced activity in vitro, but retains wild-type larvicidal toxicity. In addition, seven other mutations of charged residues result in proteins which form small or no inclusions in recombinant cells, despite being produced at levels similar to the wild-type in six out of seven cases. In most instances, the toxicity of these aberrantly expressed proteins is considerably less than the wild-type, although one (Lys124 to Ala124) results in a polypeptide with approximately threefold increased activity in vitro. A secondary structural model is proposed to explain these observations.

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Published date: 5 August 1988

Identifiers

Local EPrints ID: 425252
URI: https://eprints.soton.ac.uk/id/eprint/425252
ISSN: 0022-2836
PURE UUID: a25a13f0-1285-4529-bc85-97ab6c9ee9a1
ORCID for E. S. Ward: ORCID iD orcid.org/0000-0003-3232-7238

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Date deposited: 12 Oct 2018 16:30
Last modified: 14 Mar 2019 01:21

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Author: E. S. Ward ORCID iD
Author: D. J. Ellar
Author: C. N. Chilcott

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