Protein fiber linear dichroism for structure determination and kinetics in a low-volume, low-wavelength couette flow cell.
High-resolution structure determination of soluble globular proteins relies heavily on x-ray crystallography techniques. Such an approach is often ineffective for investigations into the structure of fibrous proteins as these proteins generally do not crystallize. Thus investigations into fibrous protein structure have relied on less direct methods such as x-ray fiber diffraction and circular dichroism. Ultraviolet linear dichroism has the potential to provide additional information on the structure of such biomolecular systems. However, existing systems are not optimized for the requirements of fibrous proteins. We have designed and built a low-volume (200 microL), low-wavelength (down to 180 nm), low-pathlength (100 microm), high-alignment flow-alignment system (couette) to perform ultraviolet linear dichroism studies on the fibers formed by a range of biomolecules. The apparatus has been tested using a number of proteins for which longer wavelength linear dichroism spectra had already been measured. The new couette cell has also been used to obtain data on two medically important protein fibers, the all-beta-sheet amyloid fibers of the Alzheimer's derived protein Abeta and the long-chain assemblies of alpha1-antitrypsin polymers.
Dafforn
Timothy R
TR
Department of Biological Sciences, University of Manchester, Manchester M13 9PT, United Kingdom. tim.dafforn@hotmail.com
Rajendra
Jacindra
J
Halsall
David J
DJ
Serpell
Louise C
LC
Rodger
Alison
A
D000199:Actins / Q000032:analysis / Q000737:chemistry*; D016229:Amyloid beta-Peptides / Q000032:analysis / Q000737:chemistry*; D002942:Circular Dichroism / Q000295:instrumentation / Q000379:methods*; D003094:Collagen / Q000032:analysis / Q000737:chemistry*; D003461:Crystallography / Q000295:instrumentation / Q000379:methods*; D007700:Kinetics; D044085:Microfluidics / Q000295:instrumentation / Q000379:methods*; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D000515:alpha 1-Antitrypsin / Q000032:analysis / Q000737:chemistry*
D000199:Actins; D016229:Amyloid beta-Peptides; D000515:alpha 1-Antitrypsin; D003094:Collagen