Positive φ-angles in proteins by nuclear magnetic resonance spectroscopy

Abstract

Summary Non-glycine residues with positive φ-angles have been identified in four proteins, barley serine proteinase inhibitor CI-2, bacterial ribonuclease (barnase) ofBacillus amyloliquefaciens, hen egg white lysozyme and a basic protein from barley seed (barwin) by use of nuclear magnetic resonance spectroscopy. By accurate measurements of the coupling constant\({}^3J_{N^N H^2 } \) and integration of the nuclear Overhauser H^N-H^α cross peak, positive φ-angles could be determined reliably to 60°±30°, in full agreement with the crystal structures for lysozyme, barnase and serine proteinase inhibitor CI-2. The work emphasizes that positive φ-angles can also occur in non-glycine residues and in the four proteins, positive φ-angles have been observed for the residue types aspartic acid, asparagine, arginine, serine, glutamine, histidine, tyrosine, tryptophan and phenylalanine. The measured\({}^3J_{N^N H^2 } \) coupling constants and the intensity of the intraresidue H^N-H^α NOEs agree well with the solution structures of three of the proteins, using the existing parametrization of the Karplus curve (Pardi, A., Billeter, M. and Wüthrich, K. (1984)J. Mol. Biol.,180, 741–751; Ludvigsen, S., Andersen, K.V. and Poulsen, F.M. (1991)J. Mol. Biol.,217, 731–736).