Open AccessKonformations‐ und Stabilitätsuntersuchungen von Polypeptiden und Proteinen mit Hilfe des H‐D‐Austausches und der Infrarot‐Spektroskopie
Author(s) -
Böhm Siegfried,
Krumbiegel Johannes,
Billwitz Heide
Publication year - 1974
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1974.tb03304.x
Subject(s) - chemistry , relative humidity , crystallography , amide , random coil , metastability , analytical chemistry (journal) , circular dichroism , physics , chromatography , organic chemistry , thermodynamics
1 Infrared spectra of nonoriented films of PGA‐Na were measured in the region of 3 μ and 6 μ both at discrete relative humidities (H 2 O and D 2 O) and under continuous variation of relative humidity between 98% and <5%. 2 The H‐D‐exchange of the different conformations of PGA‐Na films depending on the relative humidity was observed by means of the decrease of intensity of the amide A band (for nomenclature of the amide group frequencies, cf. ref. [17]). 3 Results: a) As in the case of the amide I(I′) and II(II′) bands [1,2] the position and intensity of the amide A band are dependent on conformation. Its maximum occurs at 3305 cm −1 for α‐helical structure and at 3280 cm −1 for the β‐conformation. b) The formation of α‐helix reaches a maximum value between 75 and 80% relative humidity. Above 80% relative humidity an increasing amount of random‐coil or expanded helical structures are found. c) When the relative humidity was decreased from about 75% to 69–67% (D 2 O) an unordered conformation was found to occur as the metastable intermediate state of the known α→β‐transformation. This is manifested by a strong increase of the H‐exchange rate and characteristic changes in the position and intensity of amid I(I′) and II(II′) bands. d) H‐exchange rates were found to vary drastically between the three conformations. At 25°C, the following half‐life values were determined: α‐helix (at 75–80% r.h. D 2 O) about 56 min β‐structure (at 60–74% r.h. D 2 O) about 25 h unordered structure (at 67–69% r.h. D 2 O) <2 min e) There is a hysteresis observed between the α→β and the β→α‐transformation dependent on relative humidity. Decreasing the relative humidity the α→β‐transformation starts at 67% and inceasing the relative humidity the β→α‐transformation begins at 74%. 4 The strong differences in H‐exchange rates of the three conformations are explained by different interactions between water and peptide groups.