| Author/Editor | Poklar, Nataša; Lah, Jurij; Salobir, Mateja; Maček, Peter; Vesnaver, Gorazd | |
| Title | pH and temperature-induced molten globule-like denatured states of equinatoxin II: a study by UV-melting, DSC, far- and near-UV CD spectroscopy, and ANS fluorescence | |
| Type | članek | |
| Source | Biochemistry | |
| Vol. and No. | Letnik 36, št. 47 | |
| Publication year | 1997 | |
| Volume | str. 14345-52 | |
| Language | eng | |
| Abstract | Thermal denaturation of equinatoxin II (eqTxII) in glycine buffer solutions (pH 1.1, 2.0,3.0, and 3.5) and in triple distilled water (pH 5.5-6.0) was examined by differential scanning calorimetry, UV and CD spectroscopy and fluorescence emission spectroscopy of the added hydrophobic fluorescent probe ANS. At pH 5.5-6.0 and at temperatures 60 degrees C, the protein exists in a native state characterized by a pronounced tertiary structure, a beta-rich secondary structure and a low degree of ANs-binding. At higher temperatures, it undergoes a two-state conformational transition, (deltaH degrees)HV = (deltaH degrees)DSC, into an unfolded state, which is characterized by a complete collapse of its tertiary structure and an incomplete denaturation of it secondary structure. At acidic pH, the EqTxII temperature-induced conformational transition appears at lower temperatures as non-two-state transition accompanied by the formation of an intermediate state which shows characteristics of molten globules, i.e., absence of defined tertiary structure, increase in alpha-rich secondary structure, and high affinity for ANS. At pH 2.0, the low temperature initial state of EqTxII is already partially denatured; the tertiary structure is partially discrupted, and a pronounced inequality (deltaH degrees)VH > (deltaH degrees)DSC is observed. At pH value of 1.1 and below 60 degrees C, EqTxI exists in a stable acid-denatured compact state which shows all the characteristics of a molten globule, which even at 95 degrees C is not completely denatured. According to numerous studies on the pore forming toxins, such acid-denatured compact states may contribute to the protein's ability to penetrate into biological membranes. | |
| Descriptors | CNIDARIAN VENOMS SEA ANEMONES HYDROGEN-ION CONCENTRATION TEMPERATURE PROTEIN DENATURATION BUFFERS GLYCINE PROTEIN STRUCTURE, SECONDARY PROTEIN STRUCTURE, TERTIARY CALORIMETRY, DIFFERENTIAL SCANNING SPECTROPHOTOMETRY, ULTRAVIOLET SPECTROMETRY, FLUORESCENCE |