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Localization of diphtheria toxin nuclease activity to fragment A

Lessnick, Stephen L. and Lyczak, Jeffrey B. and Bruce, Can and Lewis, Donald G. and Kim, Phillip S. and Stolowitz, Mark L. and Hood, Leroy and Wisnieski, Bernadine J. (1992) Localization of diphtheria toxin nuclease activity to fragment A. Journal of Bacteriology, 174 (6). pp. 2032-2038. ISSN 0021-9193. PMCID PMC205811.

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We describe a series of experiments that aimed to establish whether nuclease activity is actually associated with diphtheria toxin (DTx) and its A subunit (DTA), as we originally reported (M. P. Chang, R. L. Baldwin, C. Bruce, and B. J. Wisnieski, Science 246:1165-1168, 1989). Here we show that (i) trypsinization of DTx does indeed produce nucleolytically active DTA, (ii) reduction of electroeluted, unreduced, cleaved DTx (58 kDa) yields nuclease-active DTA (24 kDa), and (iii) fractionation of DTx and DTA by anion-exchange chromatography leads to coelution of nuclease activity with both forms of the toxin, even though each form elutes at a distinct salt concentration. In addition, we show that Escherichia coli-derived DTA also expresses nuclease activity. These studies confirm our initial assertion that the nuclease activity observed in DTx preparations is intrinsic to the DTA portion of DTx.

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Additional Information:© 1992 American Society for Microbiology. Received 22 July 1991; Accepted 13 January 1992. We thank A. J. Berk and D. P. Nierlich for constructive advice. Special thanks is also extended to Louis Chau for expert technical assistance. This research was supported by NIH research grant GM22240 and the UCLA Academic Senate. S.L.L. held a Short Term Training Program Grant for Excellence in Research, UCLA School of Medicine, Office of the Dean. J.B.L. is supported by USPHS training grant CA09056.
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PubMed Central ID:PMC205811
Record Number:CaltechAUTHORS:LESjbact92
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Deposited On:13 Dec 2006
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