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Assignment of the helical structure in neuropeptide Y by HPLC studies of methionine replacement analogues and 1 H‐NMR spectroscopy
Author(s) -
Rothemund Sven,
Krause Eberhard,
Beyermann Michael,
Bienert Michael,
Sykes Brian D.,
Sönnichsen Frank D.
Publication year - 1996
Publication title -
biopolymers
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.556
H-Index - 125
eISSN - 1097-0282
pISSN - 0006-3525
DOI - 10.1002/(sici)1097-0282(199608)39:2<207::aid-bip9>3.0.co;2-t
Subject(s) - chemistry , methionine sulfoxide , methionine , sulfoxide , peptide , nuclear magnetic resonance spectroscopy , circular dichroism , amphiphile , high performance liquid chromatography , dimethyl sulfoxide , helix (gastropod) , stereochemistry , crystallography , amino acid , chromatography , biochemistry , organic chemistry , ecology , snail , copolymer , biology , polymer
The HPLC retention behavior of three complete single methionine and methionine sulfoxide replacement sets of two 18‐mer model peptides and neuropeptide Y (NPY) were investigated. All peptides were prepared by multiple solid‐phase peptide synthesis. Plotting the retention time differences between methionine and methionine sulfoxide analogues vs the position of replacement shows that potentially α‐helical peptides become helical on binding during reversed‐phase high performance liquid chromatography. In the case of an amphipathic α‐helix, the retention time differences change periodically with a 3–4 repeat pattern, which allow the location of amphipathic helical structures. Replacements in nonamphipathic α‐helical domains cause local preferential binding areas and lead to sequence‐dependent retention time profiles. Methionine replacement studies of NPY suggest an unstructured or extended conformation from Tyr 1 to Ala 12 connected to a well‐defined amphipathic α‐helix from Pro 13 to Arg 35 . The assignment is confirmed by comparison of nuclear Overhauser effects based two‐dimensional 1 H‐nmr spectroscopy and utilization of the C α H shift index method in 50% trifluoroethanol/50% water. © 1996 John Wiley & Sons, Inc.