Author/Editor		Žerovnik, E; Jerala, R; Virden, R; Kroon-Žitko, L; Turk, V; Waltho, JP
Title		On the mechanism of human stefin B folding: 2. Folding from GuHCl unfolded, TFE denatured, acid denatured, and acid intermediate states
Type		članek
Source		Proteins
Vol. and No.		Letnik 32, št. 3
Publication year		1998
Volume		str. 304-13
Abstract		It has been shown that human stefin B exhibits molten globule intermediates when denatured by acid or GuHCl. In the presence of TFE, it transforms into a highly helical state. In our first study on its folding mechanism (Zerovnik et al., Proteins 32:296-303), the kinetics measured by circular dichroism (CD) and fluorescence were correlated. In the present work the kinetics of folding were monitored by tyrosine fluorescence, ANS fluorescence, and, for certain reactions, far ultraviolet (UV) CD. The folding was started from the unfolded state in 3.45 M GuHCl, the acid denatured state at pH 1.8+/-0.2, an acid molten globule intermediate I1 (pH 3.3+/-0.1, low salt), a more structured acid molten globule intermediate I2 (pH 3.3+/-0.1, 0.42 M NaCl), and the TFE state (pH 3.3+/-0.1, 42% TFE). It has been found that all denatured states, including GuHCl, TFE, acid denatured and acid molten globule intermediate I1, fold with the same kinetics, provided that the final conditions are identical. This does not apply to the second acid molten globule intermediate I2, which demonstrates a higher rate of folding by a factor of 270. Different energy of activation and pH dependence were found for folding from states I1 or I2.
Descriptors		CYSTATINS PROTEIN FOLDING ANILINO NAPHTHALENESULFONATES CIRCULAR DICHROISM FLUORESCENT DYES GUANIDINES HYDROGEN-ION CONCENTRATION KINETICS PROTEIN DENATURATION TEMPERATURE