Dynamics of the multidomain fibrinolytic protein urokinase from two-dimensional NMR.
The recent demonstrations that thrombolytic therapy with plasminogen activators can result in substantial reductions in mortality from coronary thrombosis have generated considerable interest in the properties of fibrinolytic enzymes. Examination of the primary sequence of these proteins (which include tissue plasminogen activator, plasminogen, and urokinase) reveals that each is composed of a mosaic of domains which appear to be spatially distinct and connected by short peptide linkers. There is, however, little experimental information about the three-dimensional structure of any of the proteins, although several X-ray diffraction and NMR studies of isolated domains have been reported. Here we report two-dimensional NMR spectra of intact urokinase which are remarkably well resolved for a protein of this molecular weight. This effect is a consequence of substantial independent motion between individual domains of the protein, which overcomes the broadening effects anticipated for the slow overall tumbling rate of the intact molecule. As well as having significance for the physiological role of the protein, these results provide a direct means for the comparison of structural features determined for the isolated domains with those of the intact protein and may provide a basis for proposing or evaluating models for the overall structure of fibrinolytic proteins. Preliminary results with other proteins indicate that this approach may be generally applicable to other multidomain proteins of the fibrinolytic family.
Oswald
R E
RE
Inorganic Chemistry Laboratory, University of Oxford, UK.
Bogusky
M J
MJ
Bamberger
M
M
Smith
R A
RA
Dobson
C M
CM
D004815:Epidermal Growth Factor; D005342:Fibrinolysis; D006863:Hydrogen-Ion Concentration; D046911:Macromolecular Substances; D009682:Magnetic Resonance Spectroscopy; D010446:Peptide Fragments; D012697:Serine Endopeptidases; D013329:Structure-Activity Relationship; D013816:Thermodynamics; D014568:Urokinase-Type Plasminogen Activator
D046911:Macromolecular Substances; D010446:Peptide Fragments; D004815:Epidermal Growth Factor; D012697:Serine Endopeptidases; D014568:Urokinase-Type Plasminogen Activator