Trapping the fast‐refolding state of ribonuclease A at subzero temperatures
- 29 February 1988
- journal article
- Published by Wiley in FEBS Letters
- Vol. 229 (1) , 123-126
- https://doi.org/10.1016/0014-5793(88)80810-x
Abstract
Unfolded ribonuclease A consists of a mixture of fast- and slow-refolding species. It is generally accepted that the slow-refolding states arise from isomerization of proline residues. We show that unfolding at subzero temperatures may be used to trap the fast-refolding species Uf, since the rate of proline isomerization slows down at a much faster rate than protein unfolding. The unfolding was carried out in 5 M guanidine hydrochloride; at −15°C the protein unfolding process is complete within 30 s and under these conditions there is < 1.5% proline isomerization. By using ribonuclease in which Tyr-115 was nitrated it was possible to rule out significant isomerization of Pro-114 in the observed slow-unfolding stepKeywords
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