Uniform {sup 15}N- and {sup 15}N/{sup 13}C-labeling of proteins in mammalian cells and solution structure of the amino terminal fragment of u-PA

Conference · Thu Dec 01 04:00:00 EST 1994

OSTI ID:83398

Hansen, A P; Petros, A M; Meadows, R P; Mazar, A P; Nettesheim, D G; Pederson, T M; Fesik, S W ^[1]

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1. Abbott Laboratories, Abbott Park, IL (United States)

Urokinase-type plasminogen activator (u-PA) is a 54-kDa glycoprotein that catalyzes the conversion of plasminogen to plasmin, a broad-specificity protease responsible for the degradation of fibrin clots and extracellular matrix components. The u-PA protein consists of three individual modules: a growth factor domain (GFD), a kringle, and a serine protease domain. The amino terminal fragment (ATF) includes the GFD-responsible for u-PA binding to its receptor-and the kringle domains. This protein was expressed and uniformly {sup 15}N-and {sup 15}N/{sup 13}C-labeled in mammalian cells by methods that will be described. In addition, we present the three-dimensional structure of ATF that was derived from 1299 NOE-derived distance restraints along with the {phi} angle and hydrogen bonding restraints. Although the individual domains in the structures were highly converged, the two domains are structurally independent. The overall structures of the individual domains are very similar to the structures of homologous proteins. However, important structural differences between the growth factor domain of u-PA and other homologous proteins were observed in the region that has been implicated in binding the urokinase receptor. These results may explain, in part, why other growth factors show no appreciable affinity for the urokinase receptor.

• Research Organization: Los Alamos National Lab., NM (United States)
• OSTI ID: 83398
• Report Number(s): LA--12893-C; CONF-9403228--; ON: DE95012795
• Country of Publication: United States
• Language: English

Related Subjects

40 CHEMISTRY
55 BIOLOGY AND MEDICINE
BASIC STUDIES
CARBON 13
GROWTH FACTORS
ISOTOPE APPLICATIONS
LABELLING
NITROGEN 15
PLASMINOGEN
PROTEIN STRUCTURE
PROTEINS
RECEPTORS
SERINE
UROKINASE