Related Experiment Videos
Characterization of multiple oligomeric vimentin intermediate filament units by transient electric birefringence
M Kooijman1, M Bloemendal, H van Amerongen
1Department of Biophysics, Free University, Amsterdam, The Netherlands.
Abstract:
In this work we have studied the structure of soluble vimentin units from which intermediate filaments (IFs) are built. Several oligomeric forms have been presented in the literature as IF "building blocks", but there is still no agreement on this matter. By comparing our data with various models as proposed in the literature we can favour certain models and reject others. Transient electric birefringence (TEB) measurements were performed from which information is obtained concerning electric and hydrodynamic properties of the particles under investigation. TEB decay analysis at pH 6.8 after 70 microseconds pulses (at 20 degrees C in aqueous solution) yielded three decay times: 1.1(+/- 0.3) microseconds, 4.0(+/- 1.0) microseconds and 20.0(+/- 5.0) microseconds, with amplitudes of 45% to 60%, 30% to 45%, and less than 10%, respectively. At pH 8.5 after 70 microseconds pulses, more than 90% of the TEB signal with the second decay time is obtained, while the remainder had a decay time of 15.0(+/- 4.0) microseconds. Only when the pulse duration was decreased, the fast decay time around 1 microsecond was observed, suggesting that only a minor fraction of the particles at this pH value causes such a short decay time. At both pH values, the TEB measurements indicated that, at least in part, the molecules are oriented by a permanent dipole moment. It will be shown that the shortest decay time originates from bent or flexible dimers, and the second decay time from particles with a length of 54 to 65 nm containing, at least in part, a relatively large overall dipole moment. The longest decay time is probably due to larger aggregates. These results are consistent with a model in which single dimers, antiparallel staggered tetramers and hexamers coexist. Alternatively, but less likely on the basis of literature data, a model of parallel in-register tetramers with a considerable length contribution of the head groups would fit our research.