Evidence that the A2 fragment of Shiga-like toxin type I is required for holotoxin integrity

Author:

Austin P R1,Jablonski P E1,Bohach G A1,Dunker A K1,Hovde C J1

Affiliation:

1. Department of Microbiology, Molecular Biology, and Biochemistry, University of Idaho, Moscow 83843.

Abstract

Escherichia coli Shiga-like toxin type I (SLT-I) is a potent cytotoxin consisting of an enzymatically active A subunit and a pentameric B subunit that mediates toxin binding to susceptible eukaryotic cells. Evidence that the carboxy-terminal 38 amino acids of the A subunit are involved in holotoxin 1A:5B association is presented. We compared the ability of purified recombinant SLT-I B subunit (Slt-IB) to combine in vitro with purified recombinant SLT-I A subunit (Slt-IA; full-length subunit A includes amino acids 1 to 293) and its ability to combine with purified recombinant SLT-I A1 subunit (Slt-IA1; truncated subunit A includes amino acids 1 to 255). Each mixture was analyzed for biological and physical evidence of toxin assembly. Although Slt-IA successfully combined with Slt-IB to form a molecular species similar to holotoxin that was detectable by nondenaturing polyacrylamide gel electrophoresis and immunoblotting and yielded a molecule which was cytotoxic to cultured Vero cells, Slt-IA1 did not have this ability. Slt-IA1 was 36-fold more active than Slt-IA in an in vitro protein synthesis inhibition assay. These findings suggest that the Slt-IA2 fragment is crucial for formation of SLT holotoxin and stabilizes the interaction between the A and B subunits.

Publisher

American Society for Microbiology

Subject

Infectious Diseases,Immunology,Microbiology,Parasitology

Reference49 articles.

1. Acheson D. W. K. A. Donohue-Rolfe and G. T. Keusch. 1991. The family of Shiga and Shiga-like toxins p. 415-433. In J. E. Alouf and J. H. Freer (ed.) Sourcebook of bacterial toxins. Academic Press Ltd. London.

2. Improved silver staining of plant proteins, RNA and DNA in polyacrylamide gels;Blum H.;Electrophoresis,1987

3. Inhibition of protein synthesis by Shiga toxin. Activation of the toxin and inhibition of peptide elongation;Brown J. E.;FEBS Lett.,1980

4. A system for production and rapid purification of large amounts of the Shiga toxin/Shiga-like toxin I B subunit;Calderwood S. B.;Infect. Immun.,1990

5. Nucleotide sequence of the Shiga-like toxin genes of Escherichia coli;Calderwood S. B.;Proc. Natl. Acad. Sci. USA,1987

同舟云学术

1.学者识别学者识别

2.学术分析学术分析

3.人才评估人才评估

"同舟云学术"是以全球学者为主线,采集、加工和组织学术论文而形成的新型学术文献查询和分析系统,可以对全球学者进行文献检索和人才价值评估。用户可以通过关注某些学科领域的顶尖人物而持续追踪该领域的学科进展和研究前沿。经过近期的数据扩容,当前同舟云学术共收录了国内外主流学术期刊6万余种,收集的期刊论文及会议论文总量共计约1.5亿篇,并以每天添加12000余篇中外论文的速度递增。我们也可以为用户提供个性化、定制化的学者数据。欢迎来电咨询!咨询电话:010-8811{复制后删除}0370

www.globalauthorid.com

TOP

Copyright © 2019-2024 北京同舟云网络信息技术有限公司
京公网安备11010802033243号  京ICP备18003416号-3