Insulin assembly damps conformational fluctuations: Raman analysis of amide I linewidths in native states and fibrils.
The crystal structure of insulin has been investigated in a variety of dimeric and hexameric assemblies. Interest in dynamics has been stimulated by conformational variability among crystal forms and evidence suggesting that the functional monomer undergoes a conformational change on receptor binding. Here, we employ Raman spectroscopy and Raman microscopy to investigate well-defined oligomeric species: monomeric and dimeric analogs in solution, native T(6) and R(6) hexamers in solution and corresponding polycrystalline samples. Remarkably, linewidths of Raman bands associated with the polypeptide backbone (amide I) exhibit progressive narrowing with successive self-assembly. Whereas dimerization damps fluctuations at an intermolecular beta-sheet, deconvolution of the amide I band indicates that formation of hexamers stabilizes both helical and non-helical elements. Although the structure of a monomer in solution resembles a crystallographic protomer, its encagement in a native assembly damps main-chain fluctuations. Further narrowing of a beta-sheet-specific amide I band is observed on reorganization of insulin in a cross-beta fibril. Enhanced flexibility of the native insulin monomer is in accord with molecular dynamics simulations. Such conformational fluctuations may initiate formation of an amyloidogenic nucleus and enable induced fit on receptor binding.
Dong
Jian
J
Department of Biochemistry, Case Western Reserve University School of Medicine, 10900 Euclid Avenue SOM-W427, Cleveland, OH 44106-4935, USA.
Wan
Zhuli
Z
Popov
Maxim
M
Carey
Paul R
PR
Weiss
Michael A
MA
D000818:Animals; D066298:In Vitro Techniques; D007328:Insulin / Q000737:chemistry*; D008854:Microscopy, Electron; D008954:Models, Biological; D011487:Protein Conformation; D020836:Protein Structure, Quaternary; D021122:Protein Subunits; D051381:Rats; D013059:Spectrum Analysis, Raman; D013816:Thermodynamics
D007328:Insulin; D021122:Protein Subunits