Domain characteristics of the cyanogen bromide fragment 121–316 of thermolysin
- 12 January 1983
- journal article
- research article
- Published by Wiley in International Journal of Peptide and Protein Research
- Vol. 21 (1), 49-56
- https://doi.org/10.1111/j.1399-3011.1983.tb03077.x
Abstract
The molecule of thermolysin was shown by X-ray crystallography to be composed of two structural domains of equal size comprising residues 1–157 and 158–316. In order to explore the possibility that these domains correspond to globular fragments able to refold autonomously, we have investigated the conformational and stability properties of fragment 121–316, which was obtained by limited chemical cleavage of thermolysin with cyanogen bromide. As judged by far-ultraviolet circular dichroism measurements, in aqueous solution under neutral conditions the fragment maintains a relative amount of helical structure which is comparable to that exhibited by the corresponding region in native thermolysin. The secondary structure attained by the fragment appears remarkably stable to the denaturing action of heat. By measuring the temperature dependence of the dichroic signal at 220 nm a Tm near 74d̀ was obtained. Immunodiffusion analyses indicated that the fragment recognizes and precipitates antibodies raised in rabbits using native thermolysin as immunogen. The overall conformational and immunochemical data indicate that fragment 121–316 of thermolysin is able to refold into a stable structure of native-like characteristics independently of the rest of the molecule. The results of this study complement those previously reported for fragment 206–316 (Vita, C., Fontana, A., Seeman, J.R. & Chaiken, I.M. (1979) Biochemistry18, 3023–3031).Keywords
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