title: Solution state structures of human pancreatic amylin and pramlintide.
abstract: We have employed pramlintide (prAM) as a surrogate for hAM in CD and NMR studies of the conformational preferences of the N-terminal portion of the structure in media which do not provide long-lived monomeric solutions of hAM due to its rapid conversion to preamyloid beta aggregate states. Direct comparison of hAM and prAM could be made under helix-formation-favoring conditions. On the basis of CD and NMR studies: (i) the Cys(2)-Cys(7) loop conformation has a short-span of helix (Ala(5)-Cys(7)); (ii) the extent to which this helix propagates further into the sequence is medium-dependent; a helix from Ala(5) through Ser(20) (with end fraying from His(18) onward) is observed in aqueous fluoroalcohol media; (iii) in 12+ vol.% HFIP, the amyloidogenic region of hAM forms a second helical domain (Phe(23)-Ser(29)); (iv) the two helical regions of hAM do not have any specific geometric relationship as they are connected by a flexible loop that takes different conformations and (v) although the extreme C-terminus is essential for bioactivity, it is found to be extensively randomized with conformer interconversions occurring at a much faster rate than that is observed in the remainder of the peptide sequence. Two NMR-derived structures of the 1-22 sequence fragment of hAM have been derived. The work also serves to illustrate improved methods for the NMR characterization of helices. A detailed quantitative analysis of the NOE intensities observed in aqueous HFIP revealed alternative conformations in the C-terminal portion of the common amylin helix, a region that is known to be involved in the biorecognition phenomena leading to amyloidogenesis. Even though the SNN sequence appears to be a flexible loop, the chemical shifts (and changes induced upon helix structuring) suggest some interactions between the loop and the amyloidogenic segment of hAM that occur on partial helix formation.
authors: Department of Chemistry, University of Washington, Seattle, WA 98195, USA.
authors: John R
authors: JR
authors: Cort
authors: Zhihong
authors: Z
authors: Liu
authors: Gregory M
authors: GM
authors: Lee
authors: K N L
authors: KN
authors: Huggins
authors: Susan
authors: S
authors: Janes
authors: Kathryn
authors: K
authors: Prickett
authors: Niels H
authors: NH
authors: Andersen
mesh_terms: D000595:Amino Acid Sequence; D000682:Amyloid / Q000737:chemistry*; D002021:Buffers; D002942:Circular Dichroism; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D058228:Islet Amyloid Polypeptide; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D010179:Pancreas / Q000737:chemistry*; D010446:Peptide Fragments / Q000737:chemistry*; D017433:Protein Structure, Secondary; D014867:Water / Q000737:chemistry
chemical_list: D000682:Amyloid; D002021:Buffers; D058228:Islet Amyloid Polypeptide; D010446:Peptide Fragments; D014867:Water; C105254:pramlintide