title: Hydrophobic core around tyrosine for human endothelin-1 investigated by photochemically induced dynamic nuclear polarization nuclear magnetic resonance and matrix-assisted laser desorption ionization time-of-flight mass spectrometry.
abstract: Human endothelin-1 (ET-1) is a potent cardiovascular bioactive peptide. Its activity is based on the C-terminal residues, e.g., Trp 21 in particular. Recently, we reported an NMR solution structure of ET-1, which has a C-terminal hydrophobic core around Tyr 13. This C-terminal conformation does not agree with a previously reported X-ray crystal structure. To clarify the discrepancy, we performed photo-CIDNP NMR in combination with MALDI-TOF MS. The photo-CIDNP results revealed that the Tyr 13 aromatic ring is concealed in a hydrophobic interaction. MALDI-TOF MS experiments showed this is an intramolecular interaction in monomeric form, which is also supported by sedimentation analysis and two-dimensional NMR cross-peak line shapes. Thus, we confirmed the intramolecular hydrophobic core around Tyr 13 in aqueous solution, which agrees with the solution structure. The C-terminal conformational discrepancy between the solution and crystal was caused by the intermolecular hydrogen bond between Tyr 13 of one molecule and Asp 8 of the other in a dimer-like formation of crystalline ET-1. On the other hand, we indicated that endothelin-3, another isoform of the endothelin, has an apparent self-association equilibrium under the same condition in which three tyrosines participate.
authors.affiliation: Novartis Institutes for BioMedical Research, Novartis, Ohkubo 8, Tsukuba, Ibaraki 300-2611, Japan. hiroyuki.takashima@novartis.com
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Takashima
authors.forename: Haruhiko
authors.initials: H
authors.lastname: Tamaoki
authors.forename: Naoki
authors.initials: N
authors.lastname: Teno
authors.forename: Yoshinori
authors.initials: Y
authors.lastname: Nishi
authors.forename: Susumu
authors.initials: S
authors.lastname: Uchiyama
authors.forename: Kiichi
authors.initials: K
authors.lastname: Fukui
authors.forename: Yuji
authors.initials: Y
authors.lastname: Kobayashi
mesh_terms: D000595:Amino Acid Sequence; D003460:Crystallization; D018360:Crystallography, X-Ray; D019332:Endothelin-1 / Q000737:chemistry*; D006801:Humans; D006860:Hydrogen Bonding; D057927:Hydrophobic and Hydrophilic Interactions; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D010446:Peptide Fragments / Q000737:chemistry; D010777:Photochemistry / Q000379:methods; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D012996:Solutions; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods; D013816:Thermodynamics; D014443:Tyrosine / Q000737:chemistry*; D014461:Ultracentrifugation
chemical_list: D019332:Endothelin-1; D010446:Peptide Fragments; D012996:Solutions; D014443:Tyrosine