UNSPECIFIED (1998) Differences in cytotoxicity of native and engineered RIPs can be used to assess their ability to reach the cytoplasm. BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 249 (3). pp. 637-642. ISSN 0006-291X

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Abstract

Ricin is a heterodimeric cytotoxin composed of RTB, a galactose binding lectin, and RTA, an enzymatic N-glycosidase. The toxin is endocytosed, and after intracellular routing, RTA is translocated to the cytoplasm where it inactivates ribosomes resulting in a loss of host cell protein synthesis and cell death. We show for the first time that the cytotoxicity against cultured T cells by several RTA mutants is directly proportional to the enzyme activity of RTA, suggesting this is a reliable system to measure translocation effects. Large discrepancies between cytotoxicity and enzyme action for a given pair of toxins are therefore attributable to differences in cell, binding, uptake, or membrane translocation. Fluid phase uptake and cytotoxicity of isolated RTA are essentially identical to that of the single chain toxin PAP. This important finding suggests that RTA, and the A chain of class 2 RIPs in general, has not evolved special translocation signals to complement the increased target cell binding facilitated by RTB. Experiments with the lectin RCA and with ebulin suggest those toxins have diminished cytotoxicity probably mediated by comparative deficiencies in B chain binding. Addition of a KDEL sequence to RTA increases fluid phase uptake, consistent with the notion that transport to the ER is important for cytotoxicity. Fusion of MBP or GST to the amino terminus of RTA has little effect on enzyme action or cytotoxicity. This result is not altered by protease inhibitors, suggesting the fusion proteins are probably not cleaved prior to translocation of the toxic A chain and implying that the toxins can carry large passenger proteins into the cytoplasm, an observation with interesting potential for analytical and therapeutic chemistry. (C) 1998 Academic Press.

Item Type:	Journal Article
Subjects:	Q Science > QD Chemistry Q Science > QH Natural history > QH301 Biology
Journal or Publication Title:	BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
Publisher:	ACADEMIC PRESS INC
ISSN:	0006-291X
Date:	28 August 1998
Volume:	249
Number:	3
Number of Pages:	6
Page Range:	pp. 637-642
Publication Status:	Published
URI:	http://wrap.warwick.ac.uk/id/eprint/15403

Data sourced from Thomson Reuters' Web of Knowledge

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