Analysis of the aromatic 1H NMR spectrum of chicken plasminogen kringle 4.
The intact kringle 4 domain of chicken plasminogen has been characterized by 1H NMR spectroscopy at 300 and 620 MHz in both the presence and absence of epsilon-aminocaproic acid, an antifibrinolytic drug. The study focuses on the aromatic resonances. Comparisons with spectra from human, porcine and bovine kringle 4 homologs indicates a strict conservancy of conformation, reflecting the underlying primary sequence homology, and leads to an unambiguous assignment of all the aromatic resonances, including those of Phe15 and His40 which are unique to the chicken domain. Conclusive evidence is found that the Tyr9 ring fluctuates between two states, one in which it flips fast and other in which it is severely hindered. Similarly, the Tyr64 side chain finds itself in a structurally constrained locus. The Trp62, Tyr64, and Trp72 aromatic resonances are most sensitive to ligand presence, supporting a previously reported model of the kringle 4 lysine-binding site. His40, Phe41, and Tyr74 are also perturbed by ligand indicating proximity to the site. In contrast, the Phe15 aromatic spectrum indicates a rather mobile phenyl ring which is insensitive to ligand presence, thus confirming the lesser importance of the corresponding segment within the first kringle loop in determining kringle structure and/or function.
Petros
A M
AM
Department of Chemistry, Carnegie Mellon University, Pittsburgh, Pennsylvania 15213.
Gyenes
M
M
Patthy
L
L
Llinás
M
M
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002645:Chickens; D006639:Histidine; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D010446:Peptide Fragments; D010649:Phenylalanine; D010958:Plasminogen; D012689:Sequence Homology, Nucleic Acid; D013329:Structure-Activity Relationship; D013552:Swine; D014364:Tryptophan; D014443:Tyrosine
D010446:Peptide Fragments; D014443:Tyrosine; D010649:Phenylalanine; D006639:Histidine; D014364:Tryptophan; D010958:Plasminogen