Cooperative thermal transitions of bovine and human apo-alpha-lactalbumins: evidence for a new intermediate state.
The thermal denaturation of bovine and human apo-alpha-lactalbumins at neutral pH has been studied by intrinsic protein fluorescence, circular dichroism (CD), and differential scanning microcalorimetry (DSC) methods. Apo-alpha-lactalbumin possesses a thermal transition with a midpoint about 25-30 degrees C under these conditions (pH 8.1, 10 mM borate, 1 mM EGTA), which is reflected in changes in both fluorescence emission maximum and quantum yield. However, the CD showed a decrease in ellipticity at 270 nm with a midpoint at about 10-15 degrees C, while DSC shows the transition within the region of 15-20 degrees C. The non-coincidence of transition monitored by different methods suggests the existence of an intermediate state in the course of the thermal denaturation process. This intermediate state is not the classical molten globule state which occurs at higher temperature (i.e. denatured state at these conditions) [D.A. Dolgikh, R.I. Gilmanshin, E.V. Brazhnikov, V.E. Bychkova, G.V. Semisotnov, S.Y. Venyaminov and O.B. Ptitsyn, FEBS Letters, 136 (1981) 311-315] and has physical properties intermediate between the native and molten globule states.
Veprintsev
D B
DB
Institute for Biological Instrumentation, Russian Academy of Sciences, Pushchino, Moscow region.
Permyakov
S E
SE
Permyakov
E A
EA
Rogov
V V
VV
Cawthern
K M
KM
Berliner
L J
LJ
D000818:Animals; D001059:Apoproteins / Q000737:chemistry*; D002152:Calorimetry, Differential Scanning; D002417:Cattle; D002942:Circular Dichroism; D006801:Humans; D007768:Lactalbumin / Q000737:chemistry*; D011489:Protein Denaturation; D013045:Species Specificity; D013050:Spectrometry, Fluorescence; D013696:Temperature; D013816:Thermodynamics
D001059:Apoproteins; C041966:apo-alpha-lactalbumin; D007768:Lactalbumin